Purification and biochemical properties of calmodulin from Saccharomyces cerevisiae.
Calmodulin from the yeast Saccharomyces cerevisiae was purified to complete homogeneity by hydrophobic interaction chromatography and HPLC gel filtration. The biochemical properties of the purified protein as calmodulin were examined under various criteria and its similarity and dissimilarity to other calmodulins have been described. Like other calmodulins, yeast calmodulin activated bovine phosphodiesterase and pea NAD kinase in a Ca2+-dependent manner, but its concentration for half-maximal activation was 8-10 times that of bovine calmodulin. The amino acid composition of yeast calmodulin was different from those of calmodulins from other lower eukaryotes in that it contained no tyrosine, but more leucine and had a high ratio of serine to threonine. Yeast calmodulin did not contain tryptophanyl or tyrosyl residues, so its ultraviolet spectrum reflected the absorbance of phenylalanyl residues, and had a molar absorption coefficient at 259 nm of 1900 M-1 cm-1. Ca2+ ions changed the secondary structure of yeast calmodulin, causing a 3% decrease in the alpha-helical content, unlike its effect on other calmodulins. Antibody against yeast calmodulin did not cross-react with bovine calmodulin, and antibody against bovine calmodulin did not cross-react with yeast calmodulin, presumably due to differences in the amino acid sequences of the antigenic sites. It is concluded that the molecular structure of yeast calmodulin differs from those of calmodulins from other sources, but that its Ca2+-dependent regulatory functions are highly conserved and essentially similar to those of calmodulins of higher eukaryotes.
Ohya
Y
Y
Department of Biology, Faculty of Science, University of Tokyo, Japan.
Uno
I
I
Ishikawa
T
T
Anraku
Y
Y
D000596:Amino Acids / Q000302:isolation & purification; D000906:Antibodies / Q000032:analysis; D002147:Calmodulin / Q000276:immunology / Q000302:isolation & purification*; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D007118:Immunoassay; D012441:Saccharomyces cerevisiae / Q000032:analysis*; D013056:Spectrophotometry, Ultraviolet
D000596:Amino Acids; D000906:Antibodies; D002147:Calmodulin