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Team solves structure of promising gene-targeting protein

Barry Stoddard and colleagues are first to show TAL effector structure, paving way for use in gene modification, engineering and corrective therapy

Jan. 9, 2012
Dr. Barry Stoddard

Dr. Barry Stoddard, Basic Sciences Division

Photo by Dean Forbes

Hutchinson Center researchers have solved the three-dimensional structure of a newly discovered type of gene-targeting protein shown to be useful as a DNA-targeting molecule for gene correction, gene therapy and gene modification. The findings were published online in Science Express on Jan. 5.

Using a unique form of computational and X-ray crystallographic analyses, a team of researchers led by Dr. Barry Stoddard of the Basic Sciences Division has determined the structure of a protein called a "TAL effector," which stands for "transcription-activator-like effector."

TAL effector protein

This image represents the structure of a TAL effector protein bound to its DNA target. The protein winds around the outside of the DNA double helix and "reads" the identity of each base. The ability to recognize and bind to a single unique, long DNA sequence allows the protein to act as a highly specific gene-targeting molecule.

Courtesy of the Stoddard Lab

"These proteins have a LEGO-like, modular architecture that allows them to easily be reshuffled and engineered for DNA targeting," Stoddard said. "The upcoming years will see an explosion in the development and use of TAL effectors—and more complicated molecules that are built around TAL structures—for targeted gene modification, genetic engineering and corrective gene therapy."

TAL proteins exist only in Xanthomonas, a type of gram-negative bacteria that can infect soybeans, tomatoes, peppers, rice and citrus plants, among other species. Although in nature bacteria use these proteins to target specific sites in plant DNA, they have the potential to be used in a clinical setting to help humans, Stoddard said.

Harnessing TAL effectors

"In biotechnology and medicine TAL effectors can be used by scientists to seek out and bind to DNA targets in any organism of choice, including genes in humans that contain disease-causing mutations that we might want to correct," Stoddard said, referring to a field known as targeted gene correction, which requires the development of molecules that can be delivered directly to a single DNA site. "TAL effectors have this unique capability and can be harnessed for such uses."

Since their discovery, TAL effectors have been intensely studied for gene modification applications and have been commercialized by several companies around the world. "However, until now, the lack of structure has greatly impeded the further development and improvement of TAL effectors for genetic engineering and correction," Stoddard said.

Solving the structure of the TAL effector protein allows scientists to see exactly how the protein binds to its DNA target and exactly what types of contacts it makes to the DNA in order to recognize and read each base in the DNA sequence. "By determining the structure, it is now possible to engineer the protein to work more effectively in a variety of biotech or medical applications, either by changing its DNA-targeting specificity, making the protein more stable or longer lived in cells, or by understanding how to attach additional protein modules to it that can drive desired changes in the DNA target," Stoddard said.

Contributions from collaborators

The research was conducted in collaboration with computational biologist Dr. Philip Bradley in the Public Health Sciences Division, who specializes in the computer modeling of proteins; Dr. Amanda Nga-Sze Mak, a postdoctoral fellow in Stoddard's lab; Dr. Adam Bogdanove, a professor of plant pathology and microbiology at Iowa State University, who discovered many of the properties of the TAL proteins; and Dr. Raul Cernadas, a postdoctoral research associate in Bogdanove's lab.

"The Iowa State group cloned the gene for the TAL effector, and Amanda grew crystals of the protein bound to its DNA target site and collected X-ray diffraction data," said Stoddard of the team effort. "Phil invented and pioneered many of the computational methods used in a novel manner for this study. He generated millions of models of the protein bound to its DNA, which allowed us to solve the correct structure in a very short period of time."

The National Institutes of Health, National Science Foundation, Searles Scholars Fellowship Program and Northwest Genome Engineering Consortium funded the research.



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