Identification and molecular characterization of the calmodulin-binding subunit gene (CMP1) of protein phosphatase 2B from Saccharomyces cerevisiae. An alpha-factor inducible gene.
A method has been developed for the rapid purification of yeast calmodulin in high yield. Using a 125I-labeled calmodulin SDS/PAGE gel overlay procedure with either yeast or bovine calmodulin, we show that the bovine and yeast proteins recognize the same proteins in total yeast extracts. However, yeast calmodulin does not bind to many of the proteins in vertebrate cells identified using bovine calmodulin. A lambda gt11 yeast genomic expression library was screened with yeast or bovine brain 125I-calmodulin to identify sequences derived from calmodulin binding proteins. Twelve clones were recovered, all containing a common DNA insert; all bound calmodulin in a Ca(2+)-dependent manner. The complete coding sequence was recovered and sequenced. The predicted protein sequence show greater than 50% identity to the A subunit of vertebrate protein phosphatase 2B. The gene was designated CMP1 and shown to reside on chromosome IV. Disruption or over-expression of CMP1 have no obvious phenotype; yeast appears to contain one or more CMP1-related genes. The protein product of the CMP1 gene is elevated by alpha-factor treatment, suggesting an involvement of protein phosphatase 2B in the mating response.
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Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, NY 14853.
Bretscher
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D001921:Brain / Q000378:metabolism; D002148:Calmodulin-Binding Proteins / Q000235:genetics*; D002417:Cattle; D015825:Chromosomes, Fungal; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D007527:Isoenzymes / Q000235:genetics*; D000072235:Mating Factor; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics*; D010749:Phosphoprotein Phosphatases / Q000235:genetics*; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D012689:Sequence Homology, Nucleic Acid
D002148:Calmodulin-Binding Proteins; D004271:DNA, Fungal; D007527:Isoenzymes; D010455:Peptides; D000072235:Mating Factor; D010749:Phosphoprotein Phosphatases