Casein kinase II from the yeast Yarrowia lipolytica is a heterotetramer of the form alpha alpha' beta 2. We report on the cloning and sequencing of a partial cDNA and of the complete genomic DNA coding for the catalytic alpha subunit of the casein kinase II from this yeast species. The sequence of the gene coding for this enzyme has been analyzed. No intron was found in the gene, which is present in a single copy. The deduced amino acid sequence of the gene shows high similarity with those of alpha subunit described in other species, although, uniquely, Y. lipolytica CKII alpha lacks cysteines. We find that the alpha subunit sequence of Y. lipolytica CKII is shown greater homology with the corresponding protein from S. pombe than with that from S. cerevisiae. We have analyzed CKII alpha expression and CKII alpha activity. We show that expression of this enzyme is regulated. The catalytic subunit is translated from a single mRNA, and the enzyme is present at a very low level in Y. lipolytica, as in other yeasts.