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Cell_Biology_Alberts. The lipid-linked proteins in example 5 in Figure 10–17 are made as soluble proteins in the cytosol and are subsequently anchored to the membrane by the covalent attachment of the lipid group. The proteins in example 6, however, are made as single-pass membrane proteins in the endoplasmic reticulum (ER). While still in the ER, the transmembrane segment of the protein is cleaved off and a glycosylphosphatidylinositol (GPI) anchor is added, leaving the protein bound to the noncytosolic surface of the ER membrane solely by this anchor (discussed in Chapter 12); transport vesicles eventually deliver the protein to the plasma membrane (discussed in Chapter 13).


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Cell_Biology_Alberts. By contrast to these examples, membrane-associated proteins do not extend into the hydrophobic interior of the lipid bilayer at all; they are instead bound to either face of the membrane by noncovalent interactions with other membrane proteins (Figure 10–17, examples 7 and 8). Many of the proteins of this type can be released from the membrane by relatively gentle extraction procedures, such as exposure to solutions of very high or low ionic strength or of extreme pH, which interfere with protein–protein interactions but leave the lipid bilayer intact; these proteins are often referred to as peripheral membrane proteins. Transmembrane proteins and many proteins held in the bilayer by lipid groups or hydrophobic polypeptide regions that insert into the hydrophobic core of the lipid bilayer cannot be released in these ways. lipid Anchors control the Membrane localization of Some Signaling proteins


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Cell_Biology_Alberts. lipid Anchors control the Membrane localization of Some Signaling proteins How a membrane protein is associated with the lipid bilayer reflects the function of the protein. Only transmembrane proteins can function on both sides of Figure 10–17 Various ways in which proteins associate with the lipid bilayer. Most membrane proteins are thought to extend across the bilayer as (1) a single α helix, (2) as multiple α helices, or (3) as a rolled-up β sheet (a β barrel). Some of these “single-pass” and “multipass” proteins have a covalently attached fatty acid chain inserted in the cytosolic lipid monolayer (1). Other membrane proteins are exposed at only one side of the membrane. (4) Some of these are anchored to the cytosolic surface by an amphiphilic α helix that partitions into the cytosolic monolayer of the lipid bilayer through the hydrophobic face of the helix.


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Cell_Biology_Alberts. (5) Others are attached to the bilayer solely by a covalently bound lipid chain—either a fatty acid chain or a prenyl group (see Figure 10–18)—in the cytosolic monolayer or, (6) via an oligosaccharide linker, to phosphatidylinositol in the noncytosolic monolayer—called a gpI anchor. (7, 8) Finally, membrane-associated proteins are attached to the membrane only by noncovalent interactions with other membrane proteins. the way in which the structure in (5) is formed is illustrated in Figure 10–18, while the way in which the gpI anchor shown in (6) is formed is illustrated in Figure 12–52. the details of how membrane proteins become associated with the lipid bilayer are discussed in chapter 12.


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Cell_Biology_Alberts. the bilayer or transport molecules across it. Cell-surface receptors, for example, are usually transmembrane proteins that bind signal molecules in the extracellular space and generate different intracellular signals on the opposite side of the plasma membrane. To transfer small hydrophilic molecules across a membrane, a membrane transport protein must provide a path for the molecules to cross the hydrophobic permeability barrier of the lipid bilayer; the molecular architecture of multipass transmembrane proteins (Figure 10–17, examples 2 and 3) is ideally suited for this task, as we discuss in Chapter 11.


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Cell_Biology_Alberts. Proteins that function on only one side of the lipid bilayer, by contrast, are often associated exclusively with either the lipid monolayer or a protein domain on that side. Some intracellular signaling proteins, for example, that help relay extracellular signals into the cell interior are bound to the cytosolic half of the plasma membrane by one or more covalently attached lipid groups, which can be fatty acid chains or prenyl groups (Figure 10–18). In some cases, myristic acid, a saturated 14-carbon fatty acid, is added to the N-terminal amino group of the protein during its synthesis on a ribosome. All members of the Src family of cytoplasmic protein tyrosine kinases (discussed in Chapter 15) are myristoylated in this way. Membrane attachment through a single lipid anchor is not very strong, however, and a second lipid group is often added to anchor proteins more firmly to a membrane. For most Src kinases, the second lipid modification is the attachment of palmitic acid, a


Cell_Biology_Alberts

Cell_Biology_Alberts. very strong, however, and a second lipid group is often added to anchor proteins more firmly to a membrane. For most Src kinases, the second lipid modification is the attachment of palmitic acid, a saturated 16-carbon fatty acid, to a cysteine side chain of the protein. This modification occurs in response to an extracellular signal and helps recruit the kinases to the plasma membrane. When the signaling pathway is turned off, the palmitic acid is removed, allowing the kinase to return to the cytosol. Other intracellular signaling proteins, such as the Ras family small GTPases (discussed in Chapter 15), use a combination of prenyl group and palmitic acid attachment to recruit the proteins to the plasma membrane.


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Cell_Biology_Alberts. Many proteins attach to membranes transiently. Some are classical peripheral membrane proteins that associate with membranes by regulated protein–protein interactions. Others undergo a transition from soluble to membrane protein by a conformational change that exposes a hydrophobic peptide or covalently attached lipid anchor. Many of the small GTPases of the Rab protein family that regulate intracellular membrane traffic (discussed in Chapter 13), for example, switch depending on the nucleotide that is bound to the protein. In their GDP-bound state they are soluble and free in the cytosol, whereas in their GTP-bound state their lipid anchor is exposed and tethers them to membranes. They are


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Cell_Biology_Alberts. Figure 10–18 Membrane protein attachment by a fatty acid chain or a prenyl group. the covalent attachment of either type of lipid can help localize a water-soluble protein to a membrane after its synthesis in the cytosol. (A) A fatty acid chain (myristic acid) is attached via an amide linkage to an n-terminal glycine.


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Cell_Biology_Alberts. (B) A fatty acid chain (palmitic acid) is attached via a thioester linkage to a cysteine. (c) A prenyl chain (either farnesyl or a longer geranylgeranyl chain) is attached via a thioether linkage to a cysteine residue that is initially located four residues from the protein’s c-terminus. After prenylation, the terminal three amino acids are cleaved off, and the new c-terminus is methylated before insertion of the anchor into the membrane (not shown). the structures of the lipid anchors are shown below: (D) a myristoyl anchor (derived from a 14-carbon saturated fatty acid chain), (e) a palmitoyl anchor (a 16-carbon saturated fatty acid chain), and (F) a farnesyl anchor (a 15-carbon unsaturated hydrocarbon chain).


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Cell_Biology_Alberts. Figure 10–19 a segment of a membrane-spanning polypeptide chain crossing the lipid bilayer as an α helix. Only the α-carbon backbone of the polypeptide chain is shown, with the hydrophobic amino acids in green and yellow. the polypeptide segment shown is part of the bacterial photosynthetic reaction center, the structure of which was determined by x-ray diffraction. (Based on data from J. Deisenhofer et al., Nature 318:618– 624, 1985, and h. Michel et al., EMBO J. 5:1149–1158, 1986.) membrane proteins at one moment and soluble proteins at the next. Such highly dynamic interactions greatly expand the repertoire of membrane functions. In Most transmembrane proteins, the polypeptide chain crosses the lipid Bilayer in an α-helical conformation


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Cell_Biology_Alberts. A transmembrane protein always has a unique orientation in the membrane. This reflects both the asymmetric manner in which it is inserted into the lipid bilayer in the ER during its biosynthesis (discussed in Chapter 12) and the different functions of its cytosolic and noncytosolic domains. These domains are separated by the membrane-spanning segments of the polypeptide chain, which contact the hydrophobic environment of the lipid bilayer and are composed largely of amino acids with nonpolar side chains. Because the peptide bonds themselves are polar and because water is absent, all peptide bonds in the bilayer are driven to form hydrogen bonds with one another. The hydrogen-bonding between peptide bonds is maximized if the polypeptide chain forms a regular α helix as it crosses the bilayer, and this is how most membrane-spanning segments of polypeptide chains traverse the bilayer (Figure 10–19).


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Cell_Biology_Alberts. In single-pass transmembrane proteins, the polypeptide chain crosses only once (see Figure 10–17, example 1), whereas in multipass transmembrane proteins, the polypeptide chain crosses multiple times (see Figure 10–17, example 2). An alternative way for the peptide bonds in the lipid bilayer to satisfy their hydrogen-bonding requirements is for multiple transmembrane strands of a polypeptide chain to be arranged as a βsheet that is rolled up into a cylinder (a so-called β barrel; see Figure 10–17, example 3). This protein architecture is seen in the Figure 10–20 Using hydropathy plots to porin proteins that we discuss later.


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Cell_Biology_Alberts. Progress in the x-ray crystallography of membrane proteins has enabled the determination of the three-dimensional structure of many of them. The structures confirm that it is often possible to predict from the protein’s amino acid sequence which parts of the polypeptide chain extend across the lipid bilayer. Segments containing about 20–30 amino acids, with a high degree of hydrophobicity, are long enough to span a lipid bilayer as an α helix, and they can often be identified in hydropathy plots (Figure 10–20). From such plots, it is estimated that about 30% 0 50 100 0 100 200 Rev. Biochem. 53:595–624, 1984. With amino acid number amino acid number permission from Annual reviews.) localize potential α-helical membrane-spanning segments in a polypeptide chain. the free energy needed to transfer successive segments of a polypeptide chain from a nonpolar solvent to water is calculated from the amino acid composition of each segment using data obtained from model compounds. this


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Cell_Biology_Alberts. to transfer successive segments of a polypeptide chain from a nonpolar solvent to water is calculated from the amino acid composition of each segment using data obtained from model compounds. this calculation is made for segments of a fixed size (usually around 10–20 amino acids), beginning with each successive amino acid in the chain. the “hydropathy index” of the segment is plotted on the Y axis as a function of its location in the chain. A positive value indicates that free energy is required for transfer to water (i.e., the segment is hydrophobic), and the value assigned is an index of the amount of energy needed. peaks in the hydropathy index appear at the positions of hydrophobic segments in the amino acid sequence. (A and B) hydropathy plots for two membrane proteins that are discussed later in this chapter. glycophorin (A) has a single membrane-spanning α helix, and one corresponding peak in the hydropathy plot. Bacteriorhodopsin (B) has seven membrane-spanning α helices and


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Cell_Biology_Alberts. later in this chapter. glycophorin (A) has a single membrane-spanning α helix, and one corresponding peak in the hydropathy plot. Bacteriorhodopsin (B) has seven membrane-spanning α helices and seven corresponding peaks in the hydropathy plot. (A, adapted from D. eisenberg, Annu.


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Cell_Biology_Alberts. of an organism’s proteins are transmembrane proteins, emphasizing their importance. Hydropathy plots cannot identify the membrane-spanning segments of a β barrel, as 10 amino acids or fewer are sufficient to traverse a lipid bilayer as an extended β strand and only every other amino acid side chain is hydrophobic.


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Cell_Biology_Alberts. The strong drive to maximize hydrogen-bonding in the absence of water means that a polypeptide chain that enters the lipid bilayer is likely to pass entirely through it before changing direction, since chain bending requires a loss of regular hydrogen-bonding interactions. But multipass transmembrane proteins can also contain regions that fold into the membrane from either side, squeezing into spaces between transmembrane α helices without contacting the hydrophobic core of the lipid bilayer. Because such regions interact only with other polypeptide regions, they do not need to maximize hydrogen-bonding; they can therefore have a variety of secondary structures, including helices that extend only part way across the lipid bilayer (Figure 10–21). Such regions are important for the function of some membrane proteins, including water channel and ion channel proteins, in which the regions contribute to the walls of the pores traversing the membrane and confer substrate specificity on the


Cell_Biology_Alberts

Cell_Biology_Alberts. of some membrane proteins, including water channel and ion channel proteins, in which the regions contribute to the walls of the pores traversing the membrane and confer substrate specificity on the channels, as we discuss in Chapter 11. These regions cannot be identified in hydropathy plots and are only revealed by x-ray crystallography or electron crystallography (a technique similar to x-ray diffraction but performed on two-dimensional arrays of proteins) of the protein’s three-dimensional structure.


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Cell_Biology_Alberts. transmembrane α helices Often Interact with One Another


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Cell_Biology_Alberts. The transmembrane α helices of many single-pass membrane proteins do not contribute to the folding of the protein domains on either side of the membrane. As a consequence, it is often possible to engineer cells to produce just the cytosolic or extracellular domains of these proteins as water-soluble molecules. This approach has been invaluable for studying the structure and function of these domains, especially the domains of transmembrane receptor proteins (discussed in Chapter 15). A transmembrane α helix, even in a single-pass membrane protein, however, often does more than just anchor the protein to the lipid bilayer. Many single-pass membrane proteins form homoor heterodimers that are held together by noncovalent, but strong and highly specific, interactions between the two transmembrane α helices; the sequence of the hydrophobic amino acids of these helices contains the information that directs the protein–protein interaction.


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Cell_Biology_Alberts. Similarly, the transmembrane α helices in multipass membrane proteins occupy specific positions in the folded protein structure that are determined by interactions between the neighboring helices. These interactions are crucial for the structure and function of the many channels and transporters that move molecules across cell membranes.


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Cell_Biology_Alberts. In these proteins, neighboring transmembrane helices in the folded structure of the protein shield many of the other transmembrane helices from the membrane lipids. Why, then, are these shielded helices nevertheless composed primarily of hydrophobic amino acids? The answer lies in the way in which multi-pass proteins are integrated into the membrane during their biosynthesis. As we discuss in Chapter 12, transmembrane α helices are inserted into the lipid bilayer sequentially by a protein translocator. After leaving the translocator, each helix is transiently surrounded by lipids, which requires that the helix be hydrophobic. It is only as the protein folds up into its final structure that contacts are made between adjacent helices, and protein–protein contacts replace some of the protein–lipid contacts (Figure 10–22).


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Cell_Biology_Alberts. Multipass membrane proteins that have their transmembrane segments arranged as β barrels rather than as α helices are comparatively rigid and therefore tend to form crystals readily when isolated. Thus, some of them were among the first Figure 10–21 Two short α helices in the aquaporin water channel, each of which spans only halfway through the lipid bilayer. In the plasma membrane, four monomers, one of which is shown here, form a tetramer. each monomer has a hydrophilic pore at its center, which allows water molecules to cross the membrane in single file (see Figure 11–20 and Movie 11.6). the two short colored helices are buried at an interface formed by protein–protein interactions. the mechanism by which the channel allows the passage of water molecules is discussed in more detail in chapter 11.


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Cell_Biology_Alberts. multipass membrane protein structures to be determined by x-ray crystallography. The number of β strands in a β barrel varies widely, from as few as 8 strands to as many as 22 (Figure 10–23).


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Cell_Biology_Alberts. β-barrel proteins are abundant in the outer membranes of bacteria, mitochondria, and chloroplasts. Some are pore-forming proteins, which create water-filled channels that allow selected small hydrophilic molecules to cross the membrane. The porins are well-studied examples (example 3 in Figure 10–23C). Many porin barrels are formed from a 16-strand, antiparallel β sheet rolled up into a cylindrical structure. Polar amino acid side chains line the aqueous channel on the inside, while nonpolar side chains project from the outside of the barrel to interact with the hydrophobic core of the lipid bilayer. Loops of the polypeptide chain often protrude into the lumen of the channel, narrowing it so that only certain solutes can pass. Some porins are therefore highly selective: maltoporin, for example, preferentially allows maltose and maltose oligomers to cross the outer membrane of E. coli.


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Cell_Biology_Alberts. The FepA protein is a more complex example of a β barrel transport protein (Figure 10–23D). It transports iron ions across the bacterial outer membrane. It is constructed from 22 β strands, and a large globular domain completely fills the inside of the barrel. Iron ions bind to this domain, which by an unknown mechanism moves or changes its conformation to transfer the iron across the membrane. Not all β-barrel proteins are transport proteins. Some form smaller barrels that are completely filled by amino acid side chains that project into the center of the barrel. These proteins function as receptors or enzymes (Figure 10–23A and B); the barrel serves as a rigid anchor, which holds the protein in the membrane and orients the cytosolic loops that form binding sites for specific intracellular molecules. Most multipass membrane proteins in eukaryotic cells and in the bacterial plasma membrane are constructed from transmembrane α helices. The helices


Cell_Biology_Alberts

Cell_Biology_Alberts. Most multipass membrane proteins in eukaryotic cells and in the bacterial plasma membrane are constructed from transmembrane α helices. The helices Figure 10–22 Steps in the folding of a multipass transmembrane protein. When a newly synthesized transmembrane α helix is released into the lipid bilayer, it is initially surrounded by lipid molecules. As the protein folds, contacts between the helices displace some of the lipid molecules surrounding the helices. Figure 10–23 β barrels formed from different numbers of β strands.


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Cell_Biology_Alberts. Figure 10–23 β barrels formed from different numbers of β strands. (A) the E. coli OmpA protein serves as a receptor for a bacterial virus. (B) the E. coli OMplA protein is an enzyme (a lipase) that hydrolyzes lipid molecules. the amino acids that catalyze the enzymatic reaction (shown in red) protrude from the outside surface of the barrel. (c) A porin from the bacterium Rhodobacter capsulatus forms a water-filled pore across the outer membrane. the diameter of the channel is restricted by loops (shown in blue) that protrude into the channel. (D) the E. coli FepA protein transports iron ions. the inside of the barrel is completely filled by a globular protein domain (shown in blue) that contains an iron-binding site (not shown).


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Cell_Biology_Alberts. Figure 10–24 a single-pass transmembrane protein. note that the polypeptide chain traverses the lipid bilayer as a right-handed α helix and that the oligosaccharide chains and disulfide bonds are all on the noncytosolic surface of the membrane. the sulfhydryl groups in the cytosolic domain of the protein do not normally form disulfide bonds because the reducing environment in the cytosol maintains these groups in their reduced (–Sh) form. can slide against each other, allowing conformational changes in the protein that can open and shut ion channels, transport solutes, or transduce extracellular signals into intracellular ones. In β-barrel proteins, by contrast, hydrogen bonds bind each β strand rigidly to its neighbors, making conformational changes within the wall of the barrel unlikely.


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Cell_Biology_Alberts. Most transmembrane proteins in animal cells are glycosylated. As in glycolipids, the sugar residues are added in the lumen of the ER and the Golgi apparatus (discussed in Chapters 12 and 13). For this reason, the oligosaccharide chains are always present on the noncytosolic side of the membrane. Another important difference between proteins (or parts of proteins) on the two sides of the membrane results from the reducing environment of the cytosol. This environment decreases the likelihood that intrachain or interchain disulfide (S–S) bonds will form between cysteines on the cytosolic side of membranes. These bonds form on the noncytosolic side, where they can help stabilize either the folded structure of the polypeptide chain or its association with other polypeptide chains (Figure 10–24).


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Cell_Biology_Alberts. Because the extracellular part of most plasma membrane proteins are glycosylated, carbohydrates extensively coat the surface of all eukaryotic cells. These carbohydrates occur as oligosaccharide chains covalently bound to membrane proteins (glycoproteins) and lipids (glycolipids). They also occur as the polysaccharide chains of integral membrane proteoglycan molecules. Proteoglycans, which consist of long polysaccharide chains linked covalently to a protein core, are found mainly outside the cell, as part of the extracellular matrix (discussed in Chapter 19). But, for some proteoglycans, the protein core either extends across the lipid bilayer or is attached to the bilayer by a glycosylphosphatidylinositol (GPI) anchor.


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Cell_Biology_Alberts. The terms cell coat or glycocalyx are sometimes used to describe the carbohydrate-rich zone on the cell surface. This carbohydrate layer can be visualized by various stains, such as ruthenium red (Figure 10–25A), as well as by its affinity for carbohydrate-binding proteins called lectins, which can be labeled with a fluorescent dye or some other visible marker. Although most of the sugar groups are attached to intrinsic plasma membrane molecules, the carbohydrate layer also contains both glycoproteins and proteoglycans that have been secreted into the extracellular space and then adsorbed onto the cell surface (Figure 10–25B). Many of these adsorbed macromolecules are components of the extracellular matrix, so that the boundary between the plasma membrane and the extracellular matrix is often not sharply defined. One of the many functions of the carbohydrate layer is to protect cells against mechanical and chemical damage; it also keeps various other cells at a distance, preventing


Cell_Biology_Alberts

Cell_Biology_Alberts. often not sharply defined. One of the many functions of the carbohydrate layer is to protect cells against mechanical and chemical damage; it also keeps various other cells at a distance, preventing unwanted cell–cell interactions.


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Cell_Biology_Alberts. The oligosaccharide side chains of glycoproteins and glycolipids are enormously diverse in their arrangement of sugars. Although they usually contain fewer than 15 sugars, the chains are often branched, and the sugars can be bonded together by various kinds of covalent linkages—unlike the amino acids in a polypeptide chain, which are all linked by identical peptide bonds. Even three sugars can be put together to form hundreds of different trisaccharides. Both the diversity and the exposed position of the oligosaccharides on the cell surface make them especially well suited to function in specific cell-recognition processes. As we discuss in Chapter 19, plasma-membrane-bound lectins that recognize specific oligosaccharides on cell-surface glycolipids and glycoproteins mediate a variety of transient cell–cell adhesion processes, including those occurring in lymphocyte recirculation and inflammatory responses (see Figure 19–28).


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Cell_Biology_Alberts. In general, only agents that disrupt hydrophobic associations and destroy the lipid bilayer can solubilize membrane proteins. The most useful of these for the membrane biochemist are detergents, which are small amphiphilic molecules of variable structure (Movie 10.4). Detergents are much more soluble in water than lipids. Their polar (hydrophilic) ends can be either charged (ionic), as in sodium dodecyl sulfate (SDS), or uncharged (nonionic), as in octylglucoside and Triton (Figure 10–26A). At low concentration, detergents are monomeric in solution, but when their concentration is increased above a threshold, called the critical micelle concentration (CMC), they aggregate to form micelles (Figure 10–26B–D). Above the CMC, detergent molecules rapidly diffuse in and out of micelles, keeping the concentration of monomer in the solution constant, no matter how many micelles are present. Both the CMC and the average number of detergent molecules in a micelle are characteristic properties


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Cell_Biology_Alberts. the concentration of monomer in the solution constant, no matter how many micelles are present. Both the CMC and the average number of detergent molecules in a micelle are characteristic properties of each detergent, but they also depend on the temperature, pH, and salt concentration. Detergent solutions are therefore complex systems and are difficult to study.


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Cell_Biology_Alberts. When mixed with membranes, the hydrophobic ends of detergents bind to the hydrophobic regions of the membrane proteins, where they displace lipid molecules with a collar of detergent molecules. Since the other end of the detergent Figure 10–25 The carbohydrate layer on the cell surface. (A) this electron micrograph of the surface of a lymphocyte stained with ruthenium red emphasizes the thick carbohydrate-rich layer surrounding the cell. (B) the carbohydrate layer is made up of the oligosaccharide side chains of membrane glycolipids and membrane glycoproteins and the polysaccharide chains on membrane proteoglycans. In addition, adsorbed glycoproteins, and adsorbed proteoglycans (not shown), contribute to the carbohydrate layer in many cells. note that all of the carbohydrate is on the extracellular surface of the membrane. (A, courtesy of Audrey M. glauert and g.M.W. cook.)


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Cell_Biology_Alberts. Figure 10–26 The structure and function of detergents. (A) three commonly used detergents are sodium dodecyl sulfate (SDS), an anionic detergent, and triton X-100 and β-octylglucoside, two nonionic detergents. triton X-100 is a mixture of compounds in which the region in brackets is repeated between 9 and 10 times. the hydrophobic portion of each detergent is shown in yellow, and the hydrophilic portion is shown in orange. (B) At low concentration, detergent molecules are monomeric in solution. As their concentration is increased beyond the critical micelle concentration (cMc), some of the detergent molecules form micelles. note that the concentration of detergent monomer stays constant above the cMc. (c) Because they have both polar and nonpolar ends, detergent molecules are amphiphilic; and because they are cone-shaped, they form micelles rather than bilayers (see Figure 10–7). Detergent micelles are thought to have irregular shapes, and, due to packing constraints, the hydrophobic


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Cell_Biology_Alberts. and because they are cone-shaped, they form micelles rather than bilayers (see Figure 10–7). Detergent micelles are thought to have irregular shapes, and, due to packing constraints, the hydrophobic tails are partially exposed to water. (D) the space-filling model shows the structure of a micelle composed of 20 β-octylglucoside molecules, predicted by molecular dynamics calculations. the head groups are shown in red and the hydrophobic tails in gray. (B, adapted from g. gunnarsson, B. Jönsson and h. Wennerström, J. Phys. Chem. 84:3114–3121, 1980; c, from S. Bogusz, r.M. Venable and r.W. pastor, J. Phys. Chem. B 104:5462–5470, 2000.) molecule is polar, this binding tends to bring the membrane proteins into solution as detergent–protein complexes (Figure 10–27). Usually, some lipid molecules also remain attached to the protein.


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Cell_Biology_Alberts. Strong ionic detergents, such as SDS, can solubilize even the most hydrophobic membrane proteins. This allows the proteins to be analyzed by SDS polyacrylamide-gel electrophoresis (discussed in Chapter 8), a procedure that has revolutionized the study of proteins. Such strong detergents, however, unfold (denature) proteins by binding to their internal “hydrophobic cores,” thereby rendering the proteins inactive and unusable for functional studies. Nonetheless, proteins can be readily separated and purified in their SDS-denatured form. In some cases, removal of the SDS allows the purified protein to renature, with recovery of functional activity.


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Cell_Biology_Alberts. Many membrane proteins can be solubilized and then purified in an active form by the use of mild detergents. These detergents cover the hydrophobic regions on membrane-spanning segments that become exposed after lipid removal but do not unfold the protein. If the detergent concentration of a solution of solubilized membrane proteins is reduced (by dilution, for example), membrane proteins do not remain soluble. In the presence of an excess of phospholipid molecules in such a solution, however, membrane proteins incorporate into small liposomes that form spontaneously. In this way, functionally active membrane protein systems can be reconstituted from purified components, providing a powerful means of analyzing the activities of membrane transporters, ion channels, signaling receptors, and so on (Figure 10–28). Such functional reconstitution, for example, provided proof for the hypothesis that the enzymes that make ADDITION OF PHOSPHOLIPIDS (mixed with


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Cell_Biology_Alberts. ADDITION OF PHOSPHOLIPIDS (mixed with Figure 10–27 Solubilizing a membrane protein with a mild nonionic detergent. the detergent disrupts the lipid bilayer and brings the protein into solution as protein–lipid–detergent complexes. the phospholipids in the membrane are also solubilized by the detergent, as lipid-detergent micelles. Figure 10–28 The use of mild nonionic detergents for solubilizing, purifying, and reconstituting functional membrane protein systems. In this example, functional na+-K+ pump molecules are purified and incorporated into phospholipid vesicles. this pump is present in the plasma membrane of most animal cells, where it uses the energy of Atp hydrolysis to pump na+ out of the cell and K+ in, as discussed in chapter 11. ATP (ATP synthases) use H+ gradients in mitochondrial, chloroplast, and bacterial membranes to produce ATP.


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Cell_Biology_Alberts. Membrane proteins can also be reconstituted from detergent solution into nanodiscs, which are small, uniformly sized patches of membrane that are surrounded by a belt of protein, which covers the exposed edge of the bilayer to keep the patch in solution (Figure 10–29). The belt is derived from high-density lipoproteins (HDL), which keep lipids soluble for transport in the blood. In nanodiscs the membrane protein of interest can be studied in its native lipid environment and is experimentally accessible from both sides of the bilayer, which is useful, for example, for ligand-binding experiments. Proteins contained in nanodiscs can also be analyzed by single particle electron microscopy techniques to determine their structure. By this rapidly improving technique (discussed in Chapter 9), the structure of a membrane protein can be determined to high resolution without a requirement of the protein of interest to crystallize into a regular lattice, which is often hard to achieve for


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Cell_Biology_Alberts. 9), the structure of a membrane protein can be determined to high resolution without a requirement of the protein of interest to crystallize into a regular lattice, which is often hard to achieve for membrane proteins.


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Cell_Biology_Alberts. Detergents have also played a crucial part in the purification and crystallization of membrane proteins. The development of new detergents and new expression systems that produce large quantities of membrane proteins from cDNA clones has led to a rapid increase in the number of three-dimensional structures of membrane proteins and protein complexes that are known, although they are still few compared to the known structures of water-soluble proteins and protein complexes. Bacteriorhodopsin Is a light-driven proton (h+) pump that traverses the lipid Bilayer as Seven α helices


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Cell_Biology_Alberts. Bacteriorhodopsin Is a light-driven proton (h+) pump that traverses the lipid Bilayer as Seven α helices In Chapter 11, we consider how multipass transmembrane proteins mediate the selective transport of small hydrophilic molecules across cell membranes. But a detailed understanding of how such a membrane transport protein works requires precise information about its three-dimensional structure in the bilayer. Bacteriorhodopsin was the first membrane transport protein whose structure was determined, and it has remained the prototype of many multipass membrane proteins with a similar structure.


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Cell_Biology_Alberts. The “purple membrane” of the archaeon Halobacterium salinarum is a specialized patch in the plasma membrane that contains a single species of protein molecule, bacteriorhodopsin (Figure 10–30A). The protein functions as a light-activated H+ pump that transfers H+ out of the archaeal cell. Because the bacteriorhodopsin molecules are tightly packed and arranged as a planar two-dimensional crystal (FIgure 10–30B and C), it was possible to determine their three-dimensional structure by combining electron microscopy and electron diffraction analysis—a procedure called electron crystallography, which we


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Cell_Biology_Alberts. Figure 10–29 Model of a membrane protein reconstituted into a nanodisc. When detergent is removed from a solution containing a multipass membrane protein, lipids, and a protein subunit of the high-density lipoprotein (hDl), the membrane protein becomes embedded in a small patch of lipid bilayer, which is surrounded by a belt of the hDl protein. In such nanodiscs, the hydrophobic edges of the bilayer patch are shielded by the protein belt, which renders the assembly water-soluble. mentioned earlier. This method has provided the first structural views of many membrane proteins that were found to be difficult to crystallize from detergent solutions. For bacteriorhodopsin, the structure was later confirmed and extended to very high resolution by x-ray crystallography.


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Cell_Biology_Alberts. Each bacteriorhodopsin molecule is folded into seven closely packed trans-membrane α helices and contains a single light-absorbing group, or chromophore (in this case, retinal), which gives the protein its purple color. Retinal is vitamin A in its aldehyde form and is identical to the chromophore found in rhodopsin of the photoreceptor cells of the vertebrate eye (discussed in Chapter 15). Retinal is covalently linked to a lysine side chain of the bacteriorhodopsin protein. When activated by a single photon of light, the excited chromophore changes its shape and causes a series of small conformational changes in the protein, resulting in the transfer of one H+ from the inside to the outside of the cell (Figure 10–31A). In bright light, each bacteriorhodopsin molecule can pump several hundred protons per second. The light-driven proton transfer establishes an H+ gradient across the plasma membrane, which in turn drives the production of ATP by a second protein in the cell’s plasma


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Cell_Biology_Alberts. hundred protons per second. The light-driven proton transfer establishes an H+ gradient across the plasma membrane, which in turn drives the production of ATP by a second protein in the cell’s plasma membrane. The energy stored in the H+ gradient also drives other energy-requiring processes in the cell. Thus, bacteriorhodopsin converts solar energy into a H+ gradient, which provides energy to the archaeal cell.


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Cell_Biology_Alberts. The high-resolution crystal structure of bacteriorhodopsin reveals many lipid molecules bound in specific places on the protein surface (Figure 10–31B). hydrophobic core of lipid bilayer (3 nm) Figure 10–30 Patches of purple membrane, which contain bacteriorhodopsin in the archaeon Halobacterium salinarum. (A) these archaea live in saltwater pools, where they are exposed to sunlight. they have evolved a variety of light-activated proteins, including bacteriorhodopsin, which is a light-activated h+ pump in the plasma membrane. (B) the bacteriorhodopsin molecules in the purple membrane patches are tightly packed into two-dimensional crystalline arrays. (c) Details of the molecular surface visualized by atomic force microscopy. With this technique, individual bacteriorhodopsin molecules can be seen. (D) Outline of the approximate location of the bacteriorhodopsin monomer and the individual α helices in the image shown in (c). (B–c, courtesy of Dieter Oesterhelt; D, pDB code: 2BrD.)


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Cell_Biology_Alberts. Figure 10–31 The three-dimensional structure of a bacteriorhodopsin molecule. (Movie 10.5) (A) the polypeptide chain crosses the lipid bilayer seven times as α helices. the location of the retinal chromophore (purple) and the probable pathway taken by h+ during the light-activated pumping cycle are shown. the first and key step is the passing of an h+ from the chromophore to the side chain of aspartic acid 85 (red, located next to the chromophore) that occurs upon absorption of a photon by the chromophore. Subsequently, other h+ transfers—in the numerical order indicated and utilizing the hydrophilic amino acid side chains that line a path through the membrane—complete the pumping cycle and return the enzyme to its starting state. color code: glutamic acid (orange), aspartic acid (red), arginine (blue). (B) the high-resolution crystal structure of bacteriorhodopsin shows many lipid molecules (yellow with red head groups) that are tightly bound to specific places on the surface of the


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Cell_Biology_Alberts. (blue). (B) the high-resolution crystal structure of bacteriorhodopsin shows many lipid molecules (yellow with red head groups) that are tightly bound to specific places on the surface of the protein. (A, adapted from h. luecke et al., Science 286:255–261, 1999. With permission from AAAS; B, from h. luecke et al., J. Mol. Biol. 291:899–911, 1999. With permission from Academic press.)


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Cell_Biology_Alberts. Interactions with specific lipids are thought to help stabilize many membrane proteins, which work best and sometimes crystallize more readily if some of the lipids remain bound during detergent extraction, or if specific lipids are added back to the proteins in detergent solutions. The specificity of these lipid–protein interactions helps explain why eukaryotic membranes contain such a variety of lipids, with head groups that differ in size, shape, and charge. We can think of the membrane lipids as constituting a two-dimensional solvent for the proteins in the membrane, just as water constitutes a three-dimensional solvent for proteins in an aqueous solution: some membrane proteins can function only in the presence of specific lipid head groups, just as many enzymes in aqueous solution require a particular ion for activity.


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Cell_Biology_Alberts. Bacteriorhodopsin is a member of a large superfamily of membrane proteins with similar structures but different functions. For example, rhodopsin in rod cells of the vertebrate retina and many cell-surface receptor proteins that bind extracellular signal molecules are also built from seven transmembrane α helices. These proteins function as signal transducers rather than as transporters: each responds to an extracellular signal by activating a GTP-binding protein (G protein) inside the cell and they are therefore called G-protein-coupled receptors (GPCRs), as we discuss in Chapter 15 (see Figure 15–6B). Although the structures of bacteriorhodopsins and GPCRs are strikingly similar, they show no sequence similarity and thus probably belong to two evolutionarily distant branches of an ancient protein family. A related class of membrane proteins, the channelrhodopsins that green algae use to detect light, form ion channels when they absorb a photon. When engineered so that they are


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Cell_Biology_Alberts. ancient protein family. A related class of membrane proteins, the channelrhodopsins that green algae use to detect light, form ion channels when they absorb a photon. When engineered so that they are expressed in animal brains, these proteins have become invaluable tools in neurobiology because they allow specific neurons to be stimulated experimentally by shining light on them, as we discuss in Chapter 11 (Figure 11–32).


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Cell_Biology_Alberts. Membrane proteins Often Function as large complexes Many membrane proteins function as part of multicomponent complexes, several of which have been studied by x-ray crystallography. One is a bacterial photosynthetic reaction center, which was the first membrane protein complex to be crystallized and analyzed by x-ray diffraction. In Chapter 14, we discuss how such photosynthetic complexes function to capture light energy and use it to pump H+ across the membrane. Many of the membrane protein complexes involved in photosynthesis, proton pumping, and electron transport are even larger than the photosynthetic reaction center. The enormous photosystem II complex from cyanobacteria, for example, contains 19 protein subunits and well over 60 transmembrane helices (see Figure 14–49). Membrane proteins are often arranged in large complexes, not only for harvesting various forms of energy, but also for transducing extracellular signals into intracellular ones (discussed in Chapter 15).


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Cell_Biology_Alberts. Many Membrane proteins Diffuse in the plane of the Membrane Like most membrane lipids, membrane proteins do not tumble (flip-flop) across the lipid bilayer, but they do rotate about an axis perpendicular to the plane of the bilayer (rotational diffusion). In addition, many membrane proteins are able to move laterally within the membrane (lateral diffusion). An experiment in which mouse cells were artificially fused with human cells to produce hybrid cells (heterokaryons) provided the first direct evidence that some plasma membrane proteins are mobile in the plane of the membrane. Two differently labeled antibodies were used to distinguish selected mouse and human plasma membrane proteins. Although at first the mouse and human proteins were confined to their own halves of the newly formed heterokaryon, the two sets of proteins diffused and mixed over the entire cell surface in about half an hour (Figure 10–32).


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Cell_Biology_Alberts. The lateral diffusion rates of membrane proteins can be measured by using the technique of fluorescence recovery after photobleaching (FRAP). The method usually involves marking the membrane protein of interest with a specific fluorescent group. This can be done either with a fluorescent ligand such as a fluorophore-labeled antibody that binds to the protein or with recombinant DNA technology to express the protein fused to a fluorescent protein such as green fluorescent protein (GFP) (discussed in Chapter 9). The fluorescent group is then bleached in a small area of membrane by a laser beam, and the time taken for adjacent membrane proteins carrying unbleached ligand or GFP to diffuse into the bleached area is measured (Figure 10–33). From FRAP measurements, we can estimate the diffusion coefficient for the marked cell-surface protein. The values of the diffusion coefficients for different membrane proteins in different cells are highly variable, because interactions with other


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Cell_Biology_Alberts. coefficient for the marked cell-surface protein. The values of the diffusion coefficients for different membrane proteins in different cells are highly variable, because interactions with other proteins impede the diffusion of the proteins to varying degrees. Measurements of proteins that are minimally impeded in this way indicate that cell membranes have a viscosity comparable to that of olive oil.


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Cell_Biology_Alberts. One drawback to the FRAP technique is that it monitors the movement of large populations of molecules in a relatively large area of membrane; one cannot follow individual protein molecules. If a protein fails to migrate into a bleached area, for example, one cannot tell whether the molecule is truly immobile or just restricted in its movement to a very small region of membrane—perhaps by cytoskeletal proteins. Single-particle tracking techniques overcome this problem by labeling individual membrane molecules with antibodies coupled to fluorescent dyes or tiny gold particles and tracking their movement by video microscopy. Using single-particle tracking, one can record the diffusion path of a single membrane protein molecule over time. Results from all of these techniques indicate that plasma membrane proteins differ widely in their diffusion characteristics, as we now discuss.


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Cell_Biology_Alberts. Figure 10–32 an experiment demonstrating the diffusion of proteins in the plasma membrane of mouse– human hybrid cells. In this experiment, a mouse and a human cell were fused to create a hybrid cell, which was then stained with two fluorescently labeled antibodies. One antibody (labeled with a green dye) detects mouse plasma membrane proteins, the other antibody (labeled with a red dye) detects human plasma membrane proteins. When cells were stained immediately after fusion, mouse and human plasma membrane proteins are still found in the membrane domains originating from the mouse and human cell, respectively. After a short time, however, the plasma membrane proteins diffuse over the entire cell surface and completely intermix. (From l.D. Frye and M. edidine, J. Cell Sci. 7:319–335, 1970. With permission from the company of Biologists.)


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Cell_Biology_Alberts. Figure 10–33 Measuring the rate of lateral diffusion of a membrane protein by fluorescence recovery after photobleaching. A specific protein of interest can be expressed as a fusion protein with green fluorescent protein (gFp), which is intrinsically fluorescent. the fluorescent molecules are bleached in a small area using a laser beam. the fluorescence intensity recovers as the bleached molecules diffuse away and unbleached molecules diffuse into the irradiated area (shown here in side and top views). the diffusion coefficient is calculated from a graph of the rate of recovery: the greater the diffusion coefficient of the membrane protein, the faster the recovery (Movie 10.6). cells can confine proteins and lipids to Specific Domains Within a Membrane


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Cell_Biology_Alberts. cells can confine proteins and lipids to Specific Domains Within a Membrane The recognition that biological membranes are two-dimensional fluids was a major advance in understanding membrane structure and function. It has become clear, however, that the picture of a membrane as a lipid sea in which all proteins float freely is greatly oversimplified. Most cells confine membrane proteins to specific regions in a continuous lipid bilayer. We have already discussed how bacteriorhodopsin molecules in the purple membrane of Halobacterium assemble into large two-dimensional crystals, in which individual protein molecules are relatively fixed in relationship to one another (see Figure 10–30). ATP synthase complexes in the inner mitochondrial membrane also associate into long double rows, as we discuss in Chapter 14 (see Figure 14–32). Large aggregates of this kind diffuse very slowly.


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Cell_Biology_Alberts. In epithelial cells, such as those that line the gut or the tubules of the kidney, certain plasma membrane enzymes and transport proteins are confined to the apical surface of the cells, whereas others are confined to the basal and lateral surfaces (Figure 10–34). This asymmetric distribution of membrane proteins is often essential for the function of the epithelium, as we discuss in Chapter 11 (see Figure 11–11). The lipid compositions of these two membrane domains are also different, demonstrating that epithelial cells can prevent the diffusion of lipid as well as protein molecules between the domains. The barriers set up by a specific type of intercellular junction (called a tight junction, discussed in Chapter 19; see Figure 19–18) maintain the separation of both protein and lipid molecules. Clearly, the membrane proteins that form these intercellular junctions cannot be allowed to diffuse laterally in the interacting membranes.


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Cell_Biology_Alberts. A cell can also create membrane domains without using intercellular junctions. As we already discussed, regulated protein–protein interactions in membranes are thought to create nanoscale raft domains that function in signaling and membrane trafficking. A more extreme example is seen in the mammalian spermatozoon, a single cell that consists of several structurally and functionally distinct parts covered by a continuous plasma membrane. When a sperm cell is examined by immunofluorescence microscopy with a variety of antibodies, each of which reacts with a specific cell-surface molecule, the plasma membrane is found to consist of at least three distinct domains (Figure 10–35). Some of the membrane molecules are able to diffuse freely within the confines of their own domain. The molecular nature of the “fence” that prevents the molecules from Figure 10–34 How membrane molecules can be restricted to a particular membrane domain.


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Cell_Biology_Alberts. Figure 10–34 How membrane molecules can be restricted to a particular membrane domain. In this drawing of an epithelial cell, protein A (in the apical domain of the plasma membrane) and protein B (in the basal and lateral domains) can diffuse laterally in their own domains but are prevented from entering the other domain, at least partly by the specialized cell–cell junction called a tight junction. lipid molecules in the outer (extracellular) monolayer of the plasma membrane are likewise unable to diffuse between the two domains; lipids in the inner (cytosolic) monolayer, however, are able to do so (not shown). the basal lamina is a thin mat of extracellular matrix that separates epithelial sheets from other tissues (discussed in chapter 19).


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Cell_Biology_Alberts. leaving their domain is not known. Many other cells have similar membrane fences that confine membrane protein diffusion to certain membrane domains. The plasma membrane of nerve cells, for example, contains a domain enclosing the cell body and dendrites, and another enclosing the axon; it is thought that a belt of actin filaments tightly associated with the plasma membrane at the cell-body–axon junction forms part of the barrier. Figure 10–36 shows four common ways of immobilizing specific membrane proteins through protein–protein interactions. the cortical cytoskeleton gives Membranes Mechanical Strength and restricts Membrane protein Diffusion


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Cell_Biology_Alberts. As shown in Figure 10–36B and C, a common way in which a cell restricts the lateral mobility of specific membrane proteins is to tether them to macromolecular assemblies on either side of the membrane. The characteristic biconcave shape of a red blood cell (Figure 10–37), for example, results from interactions of its plasma membrane proteins with an underlying cytoskeleton, which consists mainly of a meshwork of the filamentous protein spectrin. Spectrin is a long, thin, flexible rod about 100 nm in length. As the principal component of the red cell cytoskeleton, it maintains the structural integrity and shape of the plasma membrane, which is the red cell’s only membrane, as the cell has no nucleus or other organelles. The spectrin cytoskeleton is riveted to the membrane through various membrane proteins. The final result is a deformable, netlike meshwork that covers the entire cytosolic surface of the red cell membrane (Figure 10–38). This spectrin-based cytoskeleton enables the red


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Cell_Biology_Alberts. proteins. The final result is a deformable, netlike meshwork that covers the entire cytosolic surface of the red cell membrane (Figure 10–38). This spectrin-based cytoskeleton enables the red cell to withstand the stress on its membrane as it is forced through narrow capillaries. Mice and humans with genetic abnormalities in spectrin are anemic and have red cells that are spherical (instead of concave) and fragile; the severity of the anemia increases with the degree of spectrin deficiency.


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Cell_Biology_Alberts. Figure 10–35 Three domains in the plasma membrane of a guinea pig sperm. (A) A drawing of a guinea pig sperm. (B–D) In the three pairs of micrographs, phase-contrast micrographs are on the left, and the same cell is shown with cell-surface immunofluorescence staining on the right. Different monoclonal antibodies selectively label cell-surface molecules on (B) the anterior head, (c) the posterior head, and (D) the tail. (Micrographs courtesy of Selena carroll and Diana Myles.) Figure 10–36 Four ways of restricting the lateral mobility of specific plasma membrane proteins. (A) the proteins can self-assemble into large aggregates (as seen for bacteriorhodopsin in the purple membrane of Halobacterium salinarum); they can be tethered by interactions with assemblies of macromolecules (B) outside or (c) inside the cell; or (D) they can interact with proteins on the surface of another cell.


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Cell_Biology_Alberts. junctional complex spectrin dimer actin ankyrin band 3 glycophorin band 4.1 100 nm ˜ chain H2N HOOC ° chain fexible link between domains 106-amino-acid-long repeating domain NH2spectrin spectrin tetramer plasma membrane An analogous but much more elaborate and highly dynamic cytoskeletal network exists beneath the plasma membrane of most other cells in our body. This network, which constitutes the cortex of the cell, is rich in actin filaments, which are attached to the plasma membrane in numerous ways. The dynamic remodeling of the cortical actin network provides a driving force for many essential cell functions, including cell movement, endocytosis, and the formation of transient, mobile plasma membrane structures such as filopodia and lamellopodia band 4.1


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Cell_Biology_Alberts. Figure 10–38 The spectrin-based cytoskeleton on the cytosolic side of the human red blood cell plasma membrane. (A) the arrangement shown in the drawing has been deduced mainly from studies on the interactions of purified proteins in vitro. Spectrin heterodimers (enlarged in the drawing on the right) are linked together into a netlike meshwork by “junctional complexes” (enlarged in the drawing on the left). each spectrin heterodimer consists of two antiparallel, loosely intertwined, flexible polypeptide chains called α and β. the two spectrin chains are attached noncovalently to each other at multiple points, including at both ends. Both the α and β chains are composed largely of repeating domains. two spectrin heterodimers join end-to-end to form tetramers.


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Cell_Biology_Alberts. the junctional complexes are composed of short actin filaments (containing 13 actin monomers) and these proteins—band 4.1, adducin, and a tropomyosin molecule that probably determines the length of the actin filaments. the cytoskeleton is linked to the membrane through two transmembrane proteins—a multipass protein called band 3 and a single-pass protein called glycophorin. the spectrin tetramers bind to some band 3 proteins via ankyrin molecules, and to glycophorin and band 3 (not shown) via band 4.1 proteins.


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Cell_Biology_Alberts. (B) the electron micrograph shows the cytoskeleton on the cytosolic side of a red blood cell membrane after fixation and negative staining. the spectrin meshwork has been purposely stretched out to allow the details of its structure to be seen. In a normal cell, the meshwork shown would be much more crowded and occupy only about one-tenth of this area. (B, courtesy of t. Byers and D. Branton, Proc. Natl Acad. Sci. USA 82:6153–6157, 1985. With permission from the national Academy of Sciences.) Figure 10–37 a scanning electron micrograph of human red blood cells. the cells have a biconcave shape and lack a nucleus and other organelles (Movie 10.7). (courtesy of Bernadette chailley.) discussed in Chapter 16. The cortex of nucleated cells also contains proteins that are structurally homologous to spectrin and the other components of the red cell cytoskeleton. We discuss the cortical cytoskeleton in nucleated cells and its interactions with the plasma membrane in Chapter 16.


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Cell_Biology_Alberts. The cortical cytoskeletal network restricts diffusion of not only the plasma membrane proteins that are directly anchored to it. Because the cytoskeletal filaments are often closely apposed to the cytosolic surface of the plasma membrane, they can form mechanical barriers that obstruct the free diffusion of proteins in the membrane. These barriers partition the membrane into small domains, or corrals (Figure 10–39A), which can be either permanent, as in the sperm (see Figure 10–35), or transient. The barriers can be detected when the diffusion of individual membrane proteins is followed by high-speed, single-particle tracking. The proteins diffuse rapidly but are confined within an individual corral (Figure 10–39B); occasionally, however, thermal motions cause a few cortical filaments to detach transiently from the membrane, allowing the protein to escape into an adjacent corral.


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Cell_Biology_Alberts. The extent to which a transmembrane protein is confined within a corral depends on its association with other proteins and the size of its cytoplasmic domain; proteins with a large cytosolic domain will have a harder time passing through cytoskeletal barriers. When a cell-surface receptor binds its extracellular signal molecules, for example, large protein complexes build up on the cytosolic domain of the receptor, making it more difficult for the receptor to escape from its corral. It is thought that corralling helps concentrate such signaling complexes, increasing the speed and efficiency of the signaling process (discussed in Chapter 15).


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Cell_Biology_Alberts. Cell membranes assume many different shapes, as illustrated by the elaborate and varied structures of cell-surface protrusions and membrane-enclosed organelles in eukaryotic cells. Flat sheets, narrow tubules, round vesicles, fenestrated sheets, and pitta bread-shaped cisternae are all part of the repertoire: often, a variety of shapes will be present in different regions of the same continuous bilayer. Membrane shape is controlled dynamically, as many essential cell processes— including vesicle budding, cell movement, and cell division—require elaborate transient membrane deformations. In many cases, membrane shape is influenced by dynamic pushing and pulling forces exerted by cytoskeletal or extracellular structures, as we discuss in Chapters 13 and 16). A crucial part in producing these deformations is played by membrane-bending proteins, which control local membrane curvature. Often, cytoskeletal dynamics and membrane-bending-protein forces work together. Membrane-bending proteins


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Cell_Biology_Alberts. deformations is played by membrane-bending proteins, which control local membrane curvature. Often, cytoskeletal dynamics and membrane-bending-protein forces work together. Membrane-bending proteins attach to specific membrane regions as needed and act by one or more of three principal mechanisms: 1. Some insert hydrophobic protein domains or attached lipid anchors into one of the leaflets of a lipid bilayer. Increasing the area of only one leaflet


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Cell_Biology_Alberts. Figure 10–39 Corralling plasma membrane proteins by cortical cytoskeletal filaments. (A) the filaments are thought to provide diffusion barriers that divide the membrane into small domains, or corrals. (B) high-speed, single-particle tracking was used to follow the path of single fluorescently labeled membrane protein of one type over time. the trace shows that the individual protein molecules (the movement of each shown in a different color) diffuse within a tightly delimited membrane domain and only infrequently escape into a neighboring domain. (Adapted from A. Kusumi et al., Annu. Rev. Biophys. Biomol. Struct. 34:351–378, 2005. With permission from Annual reviews.)


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Cell_Biology_Alberts. Figure 10–40 Three ways in which membrane-bending proteins shape membranes. lipid bilayers are blue and proteins are green. (A) Bilayer without protein bound. (B) A hydrophobic region of the protein can insert as a wedge into one monolayer to pry lipid head groups apart. Such regions can either be amphiphilic helices as shown or hydrophobic hairpins. (c) the curved surface of the protein can bind to lipid head groups and deform the membrane or stabilize its curvature. (D) A protein can bind to and cluster lipids that have large head groups and thereby bend the membrane. (Adapted from W.A. prinz and J.e. hinshaw, Crit. Rev. Biochem. Mol. Biol. 44:278–291, 2009.) causes the membrane to bend (Figure 10–40B). The proteins that shape the convoluted network of narrow ER tubules are thought to work in this way. 2.


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Cell_Biology_Alberts. 2. Some membrane-bending proteins form rigid scaffolds that deform the membrane or stabilize an already bent membrane (Figure 10–40C). The coat proteins that shape the budding vesicles in intracellular transport fall into this class. 3. Some membrane-bending proteins cause particular membrane lipids to cluster together, thereby inducing membrane curvature. The ability of a lipid to induce positive or negative membrane curvature is determined by the relative cross-sectional areas of its head group and its hydrocarbon tails. For example, the large head group of phosphoinositides make these lipid molecules wedge-shaped, and their accumulation in a domain of one leaflet of a bilayer therefore induces positive curvature (Figure 10–40D). By contrast, phospholipases that remove lipid head groups produce inversely shaped lipid molecules that induce negative curvature.


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Cell_Biology_Alberts. Often, different membrane-bending proteins collaborate to achieve a particular curvature, as in shaping a budding transport vesicle, as we discuss in Chapter 13.


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Cell_Biology_Alberts. Whereas the lipid bilayer determines the basic structure of biological membranes, proteins are responsible for most membrane functions, serving as specific receptors, enzymes, transporters, and so on. Transmembrane proteins extend across the lipid bilayer. Some of these membrane proteins are single-pass proteins, in which the polypeptide chain crosses the bilayer as a single α helix. Others are multipass proteins, in which the polypeptide chain crosses the bilayer multiple times—either as a series of α helices or as a β sheet rolled up into the shape of a barrel. All proteins responsible for the transport of ions and other small water-soluble molecules through the membrane are multipass proteins. Some membrane proteins do not span the bilayer but instead are attached to either side of the membrane: some are attached to the cytosolic side by an amphipathic a helix on the protein surface or by the covalent attachment of one or more lipid chains, others are attached to the noncytosolic


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Cell_Biology_Alberts. membrane: some are attached to the cytosolic side by an amphipathic a helix on the protein surface or by the covalent attachment of one or more lipid chains, others are attached to the noncytosolic side by a GPI anchor. Some membrane-associated proteins are bound by noncovalent interactions with transmembrane proteins. In the plasma membrane of all eukaryotic cells, most of the proteins exposed on the cell surface and some of the lipid molecules in the outer lipid monolayer have oligosaccharide chains covalently attached to them. Like the lipid molecules in the bilayer, many membrane proteins are able to diffuse rapidly in the plane of the membrane. However, cells have ways of immobilizing specific membrane proteins, as well as ways of confining both membrane protein and lipid molecules to particular domains in a continuous lipid bilayer. The dynamic association of membrane-bending proteins confers on membranes their characteristic three-dimensional shapes.


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Cell_Biology_Alberts. given the highly complex lipid composition of cell membranes, what are the variations within different organelle membranes in an animal cell? What are the functional consequences of these differences, and what are the roles of the minor lipid species? Is the biophysical tendency of lipids to partition into separate phases within a lipid bilayer functionally utilized in cell membranes? If so, how is it regulated and what membrane functions does it control? how commonly do specific lipid molecules associate with membrane proteins to regulate their function? given that the structure of only a tiny fraction of all membrane proteins has been determined, what new principles of membrane protein structure remain to be discovered? Which statements are true? explain why or why not. 10–1 Although lipid molecules are free to diffuse in the plane of the bilayer, they cannot flip-flop across the bilayer unless enzyme catalysts called phospholipid translocators are present in the membrane.


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Cell_Biology_Alberts. 10–2 Whereas all the carbohydrate in the plasma membrane faces outward on the external surface of the cell, all the carbohydrate on internal membranes faces toward the cytosol. 10–3 Although membrane domains with different protein compositions are well known, there are at present no examples of membrane domains that differ in lipid composition. Discuss the following problems. 10–4 When a lipid bilayer is torn, why does it not seal itself by forming a “hemi-micelle” cap at the edges, as shown in Figure Q10–1? seal with hemi-micelle cap Figure Q10–1 A torn lipid bilayer sealed with a hypothetical “hemimicelle” cap (problem 10–4). 10–5 Margarine is made from vegetable oil by a chemical process. Do you suppose this process converts saturated fatty acids to unsaturated ones, or vice versa? Explain your answer.


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Cell_Biology_Alberts. 10–7 Monomeric single-pass transmembrane proteins span a membrane with a single α helix that has characteristic chemical properties in the region of the bilayer. Which of the three 20-amino-acid sequences listed below is the most likely candidate for such a transmembrane segment? Explain the reasons for your choice. (See back of book for one-letter amino acid code; FAMILY VW is a convenient mnemonic for hydrophobic amino acids.) 10–6 If a lipid raft is typically 70 nm in diameter and each lipid molecule has a diameter of 0.5 nm, about how many lipid molecules would there be in a lipid raft composed entirely of lipid? At a ratio of 50 lipid molecules per protein molecule (50% protein by mass), how many proteins would be in a typical raft? (Neglect the loss of lipid from the raft that would be required to accommodate the protein.) 10–8 You are studying the binding of proteins to the cytoplasmic face of cultured neuroblastoma cells and have found a method that gives a good yield of


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Cell_Biology_Alberts. would be required to accommodate the protein.) 10–8 You are studying the binding of proteins to the cytoplasmic face of cultured neuroblastoma cells and have found a method that gives a good yield of inside-out vesicles from the plasma membrane. Unfortunately, your preparations are contaminated with variable amounts of right-side-out vesicles. Nothing you have tried avoids this problem. A friend suggests that you pass your vesicles over an affinity column made of lectin coupled to solid beads. What is the point of your friend’s suggestion?


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Cell_Biology_Alberts. 10–9 Glycophorin, a protein in the plasma membrane of the red blood cell, normally exists as a homodimer that is held together entirely by interactions between its trans-membrane domains. Since transmembrane domains are hydrophobic, how is it that they can associate with one another so specifically? 10–10 Three mechanisms by which membrane-binding proteins bend a membrane are illustrated in Figure Q10–2A, B, and C. As shown, each of these cytosolic membrane-bending proteins would induce an invagination of the plasma membrane. Could similar kinds of cytosolic proteins induce a protrusion of the plasma membrane (Figure Q10–2D)? Which ones? Explain how they might work.


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Cell_Biology_Alberts. Figure Q10–2 Bending of the plasma membrane by cytosolic proteins (problem 10–10). (A) Insertion of a protein “finger” into the cytosolic leaflet of the membrane. (B) Binding of lipids to the curved surface of a membrane-binding protein. (c) Binding of membrane proteins to membrane lipids with large head groups. (D) A segment of the plasma membrane showing a protrusion. Bretscher MS (1973) Membrane structure: some general principles. Science 181, 622–629. edidin M (2003) lipids on the frontier: a century of cell-membrane bilayers. Nat. Rev. Mol. Cell Biol. 4, 414–418. goñi FM (2014) the basic structure and dynamics of cell membranes: an update of the Singer-nicolson model. Biochim. Biophys. Acta 1838, 1467–1476. lipowsky r & Sackmann e (eds) (1995) Structure and Dynamics of Membranes. Amsterdam: elsevier. Singer SJ & nicolson gl (1972) the fluid mosaic model of the structure of cell membranes. Science 175, 720–731.


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Cell_Biology_Alberts. Singer SJ & nicolson gl (1972) the fluid mosaic model of the structure of cell membranes. Science 175, 720–731. tanford c (1980) the hydrophobic effect: Formation of Micelles and Biological Membranes. new York: Wiley. The Lipid bilayer Bevers eM, comfurius p, Dekkers DW & zwaal rF (1999) lipid translocation across the plasma membrane of mammalian cells. Biochim. Biophys. Acta 1439, 317–330. Brügger B (2014) lipidomics: analysis of the lipid composition of cells and subcellular organelles by electrospray ionization mass spectrometry. Annu. Rev. Biochem. 83, 79–98. contreras FX, Sánchez-Magraner l, Alonso A & goñi FM (2010) transbilayer (flip-flop) lipid motion and lipid scrambling in membranes. FEBS Lett. 584, 1779–1786. hakomori SI (2002) the glycosynapse. Proc. Natl Acad. Sci. USA 99, 225–232. Ichikawa S & hirabayashi Y (1998) glucosylceramide synthase and glycosphingolipid synthesis. Trends Cell Biol. 8, 198–202.


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Cell_Biology_Alberts. Ichikawa S & hirabayashi Y (1998) glucosylceramide synthase and glycosphingolipid synthesis. Trends Cell Biol. 8, 198–202. Klose c, Surma MA & Simons K (2013) Organellar lipidomics— background and perspectives. Curr. Opin. Cell Biol. 25, 406–413. Kornberg rD & Mcconnell hM (1971) lateral diffusion of phospholipids in a vesicle membrane. Proc. Natl Acad. Sci. USA 68, 2564–2568. lingwood D & Simons K (2010) lipid rafts as a membrane-organizing principle. Science 327, 46–50. Mansilla Mc, cybulski le, Albanesi D & de Mendoza D (2004) control of membrane lipid fluidity by molecular thermosensors. J. Bacteriol. 186, 6681–6688. Maxfield Fr & van Meer g (2010) cholesterol, the central lipid of mammalian cells. Curr. Opin. Cell Biol. 22, 422–429. Mcconnell hM & radhakrishnan A (2003) condensed complexes of cholesterol and phospholipids. Biochim. Biophys. Acta 1610, 159–173. pomorski t & Menon AK (2006) lipid flippases and their biological functions. Cell. Mol. Life Sci. 63, 2908–2921.


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Cell_Biology_Alberts. pomorski t & Menon AK (2006) lipid flippases and their biological functions. Cell. Mol. Life Sci. 63, 2908–2921. rothman Je & lenard J (1977) Membrane asymmetry. Science 195, 743–753. Walther tc & Farese rV Jr (2012) lipid droplets and cellular lipid metabolism. Annu. Rev. Biochem. 81, 687–714. Bennett V & Baines AJ (2001) Spectrin and ankyrin-based pathways: metazoan inventions for integrating cells into tissues. Physiol. Rev. 81, 1353–1392. Bijlmakers MJ & Marsh M (2003) the on-off story of protein palmitoylation. Trends Cell Biol. 13, 32–42. Branden c & tooze J (1999) Introduction to protein Structure, 2nd ed. new York: garland Science. Bretscher MS & raff Mc (1975) Mammalian plasma membranes. Nature 258, 43–49. Buchanan SK (1999) Beta-barrel proteins from bacterial outer membranes: structure, function and refolding. Curr. Opin. Struct. Biol. 9, 455–461.


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Cell_Biology_Alberts. Buchanan SK (1999) Beta-barrel proteins from bacterial outer membranes: structure, function and refolding. Curr. Opin. Struct. Biol. 9, 455–461. chen Y, lagerholm Bc, Yang B & Jacobson K (2006) Methods to measure the lateral diffusion of membrane lipids and proteins. Methods 39, 147–153. curran Ar & engelman DM (2003) Sequence motifs, polar interactions and conformational changes in helical membrane proteins. Curr. Opin. Struct. Biol. 13, 412–417. Deisenhofer J & Michel h (1991) Structures of bacterial photosynthetic reaction centers. Annu. Rev. Cell Biol. 7, 1–23. Drickamer K & taylor Me (1993) Biology of animal lectins. Annu. Rev. Cell Biol. 9, 237–264. Drickamer K & taylor Me (1998) evolving views of protein glycosylation. Trends Biochem. Sci. 23, 321–324. Frye lD & edidin M (1970) the rapid intermixing of cell surface antigens after formation of mouse-human heterokaryons. J. Cell Sci. 7, 319–335.


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Cell_Biology_Alberts. Frye lD & edidin M (1970) the rapid intermixing of cell surface antigens after formation of mouse-human heterokaryons. J. Cell Sci. 7, 319–335. helenius A & Simons K (1975) Solubilization of membranes by detergents. Biochim. Biophys. Acta 415, 29–79. henderson r & Unwin pn (1975) three-dimensional model of purple membrane obtained by electron microscopy. Nature 257, 28–32. Kyte J & Doolittle rF (1982) A simple method for displaying the hydropathic character of a protein. J. Mol. Biol. 157, 105–132. lee Ag (2003) lipid-protein interactions in biological membranes: a structural perspective. Biochim. Biophys. Acta 1612, 1–40. Marchesi Vt, Furthmayr h & tomita M (1976) the red cell membrane. Annu. Rev. Biochem. 45, 667–698. nakada c, ritchie K, Oba Y et al. (2003) Accumulation of anchored proteins forms membrane diffusion barriers during neuronal polarization. Nat. Cell Biol. 5, 626–632.


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Cell_Biology_Alberts. nakada c, ritchie K, Oba Y et al. (2003) Accumulation of anchored proteins forms membrane diffusion barriers during neuronal polarization. Nat. Cell Biol. 5, 626–632. Oesterhelt D (1998) the structure and mechanism of the family of retinal proteins from halophilic archaea. Curr. Opin. Struct. Biol. 8, 489–500. popot J-l (2010) Amphipols, nanodiscs, and fluorinated surfactants: three nonconventional approaches to studying membrane proteins in aqueous solution. Annu. Rev. Biochem. 79, 737–775. prinz WA & hinshaw Je (2009) Membrane-bending proteins. Crit. Rev. Biochem. Mol. Biol. 44, 278–291. rao M & Mayor S (2014) Active organization of membrane constituents in living cells. Curr. Opin. Cell Biol. 29, 126–132. reig n & van der goot Fg (2006) About lipids and toxins. FEBS Lett. 580, 5572–5579. Sharon n & lis h (2004) history of lectins: from hemagglutinins to biological recognition molecules. Glycobiology 14, 53r–62r.


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Cell_Biology_Alberts. Sharon n & lis h (2004) history of lectins: from hemagglutinins to biological recognition molecules. Glycobiology 14, 53r–62r. Sheetz Mp (2001) cell control by membrane-cytoskeleton adhesion. Nat. Rev. Mol. Cell Biol. 2, 392–396. Shibata Y, hu J, Kozlov MM & rapoport tA (2009) Mechanisms shaping the membranes of cellular organelles. Annu. Rev. Cell Dev. Biol. 25, 329–354. Steck tl (1974) the organization of proteins in the human red blood cell membrane. A review. J. Cell Biol. 62, 1–19. Subramaniam S (1999) the structure of bacteriorhodopsin: an emerging consensus. Curr. Opin. Struct. Biol. 9, 462–468. Viel A & Branton D (1996) Spectrin: on the path from structure to function. Curr. Opin. Cell Biol. 8, 49–55. Vinothkumar Kr & henderson r (2010) Structures of membrane proteins. Q. Rev. Biophys. 43, 65–158. von heijne g (2011) Membrane proteins: from bench to bits. Biochem. Soc. Trans. 39, 747–750.


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Cell_Biology_Alberts. Vinothkumar Kr & henderson r (2010) Structures of membrane proteins. Q. Rev. Biophys. 43, 65–158. von heijne g (2011) Membrane proteins: from bench to bits. Biochem. Soc. Trans. 39, 747–750. White Sh & Wimley Wc (1999) Membrane protein folding and stability: physical principles. Annu. Rev. Biophys. Biomol. Struct. 28, 319–365. Membrane Transport of Small Molecules and the Electrical Properties of Membranes 11


Cell_Biology_Alberts

Cell_Biology_Alberts. Because of its hydrophobic interior, the lipid bilayer of cell membranes restricts the passage of most polar molecules. This barrier function allows the cell to maintain concentrations of solutes in its cytosol that differ from those in the extracellular fluid and in each of the intracellular membrane-enclosed compartments. To benefit from this barrier, however, cells have had to evolve ways of transferring specific water-soluble molecules and ions across their membranes in order to ingest essential nutrients, excrete metabolic waste products, and regulate intracellular ion concentrations. Cells use specialized membrane transport proteins to accomplish this goal. The importance of such small molecule transport is reflected in the large number of genes in all organisms that code for the transmembrane transport proteins involved, which make up 15–30% of the membrane proteins in all cells. Some mammalian cells, such as nerve and kidney cells, devote up to two-thirds of their total


Cell_Biology_Alberts

Cell_Biology_Alberts. the transmembrane transport proteins involved, which make up 15–30% of the membrane proteins in all cells. Some mammalian cells, such as nerve and kidney cells, devote up to two-thirds of their total metabolic energy consumption to such transport processes.


Cell_Biology_Alberts

Cell_Biology_Alberts. Cells can also transfer macromolecules and even large particles across their membranes, but the mechanisms involved in most of these cases differ from those used for transferring small molecules, and they are discussed in Chapters 12 and 13.


Cell_Biology_Alberts

Cell_Biology_Alberts. We begin this chapter by describing some general principles of how small water-soluble molecules traverse cell membranes. We then consider, in turn, the two main classes of membrane proteins that mediate this transmembrane traffic: transporters, which undergo sequential conformational changes to transport specific small molecules across membranes, and channels, which form narrow pores, allowing passive transmembrane movement, primarily of water and small inorganic ions. Transporters can be coupled to a source of energy to catalyze active transport, which together with selective passive permeability, creates large differences in the composition of the cytosol compared with that of either the extracellular fluid (Table 11–1) or the fluid within membrane-enclosed organelles. By generating inorganic ion-concentration differences across the lipid bilayer, cell membranes can store potential energy in the form of electrochemical gradients, which drive various transport processes, convey


Cell_Biology_Alberts

Cell_Biology_Alberts. inorganic ion-concentration differences across the lipid bilayer, cell membranes can store potential energy in the form of electrochemical gradients, which drive various transport processes, convey electrical signals in electrically excitable cells, and (in mitochondria, chloroplasts, and bacteria) make most of the cell’s ATP. We focus our discussion mainly on transport across the plasma membrane, but similar mechanisms operate across the other membranes of the eukaryotic cell, as discussed in later chapters.


Cell_Biology_Alberts

Cell_Biology_Alberts. In the last part of the chapter, we concentrate mainly on the functions of ion channels in neurons (nerve cells). In these cells, channel proteins perform at their highest level of sophistication, enabling networks of neurons to carry out all the astonishing feats your brain is capable of. We begin this section by describing the permeability properties of protein-free, synthetic lipid bilayers. We then introduce some of the terms used to describe the various forms of membrane transport and some strategies for characterizing the proteins and processes involved. Protein-Free Lipid Bilayers Are Impermeable to Ions


Cell_Biology_Alberts

Cell_Biology_Alberts. Protein-Free Lipid Bilayers Are Impermeable to Ions Given enough time, virtually any molecule will diffuse across a protein-free lipid bilayer down its concentration gradient. The rate of diffusion, however, varies enormously, depending partly on the size of the molecule but mostly on its relative hydrophobicity (solubility in oil). In general, the smaller the molecule and the more hydrophobic, or nonpolar, it is, the more easily it will diffuse across a lipid bilayer. Small nonpolar molecules, such as O2 and CO2, readily dissolve in lipid bilayers and therefore diffuse rapidly across them. Small uncharged polar molecules, such as water or urea, also diffuse across a bilayer, albeit much more slowly (Figure 11–1 and see Movie 10.3). By contrast, lipid bilayers are essentially impermeable to charged molecules (ions), no matter how small: the charge and high degree of hydration of such molecules prevents them from entering the hydrocarbon phase of the bilayer (Figure 11–2).


Cell_Biology_Alberts

Cell_Biology_Alberts. There Are Two Main Classes of Membrane Transport Proteins: Transporters and Channels


Cell_Biology_Alberts

Cell_Biology_Alberts. Like synthetic lipid bilayers, cell membranes allow small nonpolar molecules to permeate by diffusion. Cell membranes, however, also have to allow the passage of various polar molecules, such as ions, sugars, amino acids, nucleotides, water, and many cell metabolites that cross synthetic lipid bilayers only very slowly. Special membrane transport proteins transfer such solutes across cell membranes. These proteins occur in many forms and in all types of biological membranes. Each protein often transports only a specific molecular species or sometimes a class of molecules (such as ions, sugars, or amino acids). Studies in the 1950s found that bacteria with a single-gene mutation were unable to transport sugars across their plasma membrane, thereby demonstrating the specificity of membrane transport proteins. We now know that humans with similar mutations suffer from various inherited diseases that hinder the transport of a specific solute or solute class in the kidney, intestine, or


Cell_Biology_Alberts

Cell_Biology_Alberts. transport proteins. We now know that humans with similar mutations suffer from various inherited diseases that hinder the transport of a specific solute or solute class in the kidney, intestine, or other cell type. Individuals with the inherited disease cystinuria, for example, cannot transport certain amino acids (including cystine, the disulfide-linked dimer of cysteine) from either the urine or the intestine into the


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 11–1 The relative permeability of a synthetic lipid bilayer to different classes of molecules. The smaller the molecule and, more importantly, the less strongly it associates with water, the more rapidly the molecule diffuses across the bilayer. Figure 11–2 Permeability coefficients for the passage of various molecules through synthetic lipid bilayers. The rate of flow of a solute across the bilayer is directly proportional to the difference in its concentration on the two sides of the membrane. Multiplying this concentration difference (in mol/cm3) by the permeability coefficient (in cm/sec) gives the flow of solute in moles per second per square centimeter of bilayer. A concentration difference of tryptophan of 10–4 mol/cm3 (10–4 mol / 10–3 L = 0.1 M), for example, would cause a flow of 10–4 mol/cm3 × 10–7 cm/sec = 10–11 mol/sec through 1 cm2 of bilayer, or 6 × 104 molecules/sec through 1 μm2 of bilayer.


Cell_Biology_Alberts

Cell_Biology_Alberts. blood; the resulting accumulation of cystine in the urine leads to the formation of cystine stones in the kidneys. All membrane transport proteins that have been studied in detail are multi-pass transmembrane proteins—that is, their polypeptide chains traverse the lipid bilayer multiple times. By forming a protein-lined pathway across the membrane, these proteins enable specific hydrophilic solutes to cross the membrane without coming into direct contact with the hydrophobic interior of the lipid bilayer.


Cell_Biology_Alberts

Cell_Biology_Alberts. Transporters and channels are the two major classes of membrane transport proteins (Figure 11–3). Transporters (also called carriers, or permeases) bind the specific solute to be transported and undergo a series of conformational changes that alternately expose solute-binding sites on one side of the membrane and then on the other to transfer the solute across it. Channels, by contrast, interact with the solute to be transported much more weakly. They form continuous pores that extend across the lipid bilayer. When open, these pores allow specific solutes (such as inorganic ions of appropriate size and charge and in some cases small molecules, including water, glycerol, and ammonia) to pass through them and thereby cross the membrane. Not surprisingly, transport through channels occurs at a much faster rate than transport mediated by transporters. Although water can slowly diffuse across synthetic lipid bilayers, cells use dedicated channel proteins (called water channels, or


Cell_Biology_Alberts

Cell_Biology_Alberts. at a much faster rate than transport mediated by transporters. Although water can slowly diffuse across synthetic lipid bilayers, cells use dedicated channel proteins (called water channels, or aquaporins) that greatly increase the permeability of their membranes to water, as we discuss later.


Cell_Biology_Alberts

Cell_Biology_Alberts. Active Transport Is Mediated by Transporters Coupled to an Energy Source


Cell_Biology_Alberts

Cell_Biology_Alberts. All channels and many transporters allow solutes to cross the membrane only passively (“downhill”), a process called passive transport. In the case of transport of a single uncharged molecule, the difference in the concentration on the two sides of the membrane—its concentration gradient—drives passive transport and determines its direction (Figure 11–4A). If the solute carries a net charge, however, both its concentration gradient and the electrical potential difference across the membrane, the membrane potential, influence its transport. The concentration gradient and the electrical gradient combine to form a net driving force, the electrochemical gradient, for each charged solute (Figure 11–4B). We discuss electrochemical gradients in more detail later and in Chapter 14. In fact, almost all plasma membranes have an electrical potential (i.e., a voltage) across them, with the inside usually negative with respect to the outside. This potential favors the entry of positively charged


Cell_Biology_Alberts

Cell_Biology_Alberts. all plasma membranes have an electrical potential (i.e., a voltage) across them, with the inside usually negative with respect to the outside. This potential favors the entry of positively charged ions into the cell but opposes the entry of negatively charged ions (see Figure 11–4B); it also opposes the efflux of positively charged ions.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 11–3 Transporters and channel proteins. (A) A transporter alternates between two conformations, so that the solute-binding site is sequentially accessible on one side of the bilayer and then on the other. (B) In contrast, a channel protein forms a pore across the bilayer through which specific solutes can passively diffuse. concentration electrochemical gradient with gradient (with no a membrane potential As shown in Figure 11–4A, in addition to passive transport, cells need to be able to actively pump certain solutes across the membrane “uphill,” against their electrochemical gradients. Such active transport is mediated by transporters whose pumping activity is directional because it is tightly coupled to a source of metabolic energy, such as an ion gradient or ATP hydrolysis, as discussed later. Transmembrane movement of small molecules mediated by transporters can be either active or passive, whereas that mediated by channels is always passive (see Figure 11–4A).


Cell_Biology_Alberts

Cell_Biology_Alberts. Lipid bilayers are virtually impermeable to most polar molecules. To transport small water-soluble molecules into or out of cells or intracellular membrane-enclosed compartments, cell membranes contain various membrane transport proteins, each of which is responsible for transferring a particular solute or class of solutes across the membrane. There are two classes of membrane transport proteins—transporters and channels. Both form protein pathways across the lipid bilayer. Whereas transmembrane movement mediated by transporters can be either active or passive, solute flow through channel proteins is always passive. Both active and passive ion transport is influenced by the ion’s concentration gradient and the membrane potential—that is, its electrochemical gradient.


Cell_Biology_Alberts

Cell_Biology_Alberts. The process by which a transporter transfers a solute molecule across the lipid bilayer resembles an enzyme–substrate reaction, and in many ways transporters behave like enzymes. By contrast to ordinary enzyme–substrate reactions, however, the transporter does not modify the transported solute but instead delivers it unchanged to the other side of the membrane. Each type of transporter has one or more specific binding sites for its solute (substrate). It transfers the solute across the lipid bilayer by undergoing reversible


Cell_Biology_Alberts

Cell_Biology_Alberts. Each type of transporter has one or more specific binding sites for its solute (substrate). It transfers the solute across the lipid bilayer by undergoing reversible Figure 11–4 Different forms of membrane transport and the influence of the membrane. Passive transport down a concentration gradient (or an electrochemical gradient—see B below) occurs spontaneously, by diffusion, either through the lipid bilayer directly or through channels or passive transporters. By contrast, active transport requires an input of metabolic energy and is always mediated by transporters that pump the solute against its concentration or electrochemical gradient. (B) The electrochemical gradient of a charged solute (an ion) affects its transport. This gradient combines the membrane potential and the concentration gradient of the solute. The electrical and chemical gradients can work additively to increase the driving force on an ion across the membrane (middle) or can work against each other (right).


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 11–5 a model of how a conformational change in a transporter mediates the passive movement of a solute. The transporter is shown in three conformational states: in the outward- open state, the binding sites for solute are exposed on the outside; in the occluded state, the same sites are not accessible from either side; and in the inward-open state, the sites are exposed on the inside. The transitions between the states occur randomly. They are completely reversible and do not depend on whether the solute-binding site is occupied. Therefore, if the solute concentration is higher on the outside of the bilayer, more solute binds to the transporter in the outward-open conformation than in the inward-open conformation, and there is a net transport of solute down its concentration gradient (or, if the solute is an ion, down its electrochemical gradient).


Cell_Biology_Alberts

Cell_Biology_Alberts. conformational changes that alternately expose the solute-binding site first on one side of the membrane and then on the other—but never on both sides at the same time. The transition occurs through an intermediate state in which the solute is inaccessible, or occluded, from either side of the membrane (Figure 11–5). When the transporter is saturated (that is, when all solute-binding sites are occupied), the rate of transport is maximal. This rate, referred to as Vmax (V for velocity), is characteristic of the specific carrier. Vmax measures the rate at which the carrier can flip between its conformational states. In addition, each transporter has a characteristic affinity for its solute, reflected in the Km of the reaction, which is equal to the concentration of solute when the transport rate is half its maximum value (Figure 11–6). As with enzymes, the binding of solute can be blocked by either competitive inhibitors (which compete for the same binding site and may or may not be


Cell_Biology_Alberts

Cell_Biology_Alberts. rate is half its maximum value (Figure 11–6). As with enzymes, the binding of solute can be blocked by either competitive inhibitors (which compete for the same binding site and may or may not be transported) or noncompetitive inhibitors (which bind elsewhere and alter the structure of the transporter).


Cell_Biology_Alberts

Cell_Biology_Alberts. As we discuss shortly, it requires only a relatively minor modification of the model shown in Figure 11–5 to link a transporter to a source of energy in order to pump a solute uphill against its electrochemical gradient. Cells carry out such active transport in three main ways (Figure 11–7): 1. Coupled transporters harness the energy stored in concentration gradients to couple the uphill transport of one solute across the membrane to the downhill transport of another. 2. ATP-driven pumps couple uphill transport to the hydrolysis of ATP. 3. Lightor redox-driven pumps, which are known in bacteria, archaea, mitochondria, and chloroplasts, couple uphill transport to an input of energy from light, as with bacteriorhodopsin (discussed in Chapter 10), or from a rate of transport redox reaction, as with cytochrome c oxidase (discussed in Chapter 14).


Cell_Biology_Alberts

Cell_Biology_Alberts. Amino acid sequence and three-dimensional structure comparisons suggest that, in many cases, there are strong similarities in structure between transporters that mediate active transport and those that mediate passive transport. Some bacterial transporters, for example, that use the energy stored in the H+ gradient across the plasma membrane to drive the active uptake of various sugars are structurally similar to the transporters that mediate passive glucose transport into most animal cells. This suggests an evolutionary relationship between various transporters. Given the importance of small metabolites and sugars as energy sources, it is not surprising that the superfamily of transporters is an ancient one.


Cell_Biology_Alberts

Cell_Biology_Alberts. We begin our discussion of active membrane transport by considering a class of coupled transporters that are driven by ion concentration gradients. These proteins have a crucial role in the transport of small metabolites across membranes in all cells. We then discuss ATP-driven pumps, including the Na+-K+ pump that is found in the plasma membrane of most animal cells. Examples of the third class of active transport—lightor redox-driven pumps—are discussed in Chapter 14. Some transporters simply passively mediate the movement of a single solute from one side of the membrane to the other at a rate determined by their Vmax and