Kinetics and regulation of two catalytic subunits of cAMP-dependent protein kinase from Aplysia californica.
CAPL-A1 and CAPL-A2, two catalytic subunits of Aplysia cAMP-dependent protein kinase, are encoded by mRNAs generated by alternative splicing of transcripts of a gene that contains two mutually exclusive exon cassettes. The subunits are identical except for amino acids 142-183 of the 352 residues, which differ at 10 of 42 positions. CAPL-A1 and CAPL-A2 have now been expressed in insect cells and purified to homogeneity. The subunits differ in their catalytic properties, which have been determined with a series of synthetic peptide substrates. For example, kcat and Km values for the peptide LRRASLG (kemptide) are 42 s-1 and 36 microM and 28 s-1 and 17 microM for CAPL-A1 and CAPL-A2, respectively. CAPL-A1 and CAPL-A2 have different substrate specificities. For example, (kcat/Km)peptide-T/(kcat/Km)kemptide is 9.1 x 10(-3) for CAPL-A1 and 15 x 10(-3) for CAPL-A2, where peptide-T is the kemptide homologue LRRATLG. The subunits also differ in regulation as determined by their interactions with a purified type I regulatory subunit, which has an IC50 for CAPL-A1 that is 3.5 times higher than the IC50 for CAPL-A2. These modest differences reinforce accumulating evidence that the physiological state of a cell depends upon a spectrum of protein kinases with overlapping substrate specificities and regulatory properties.
Cheley
S
S
Worcester Foundation for Experimental Biology, Shrewsbury, Massachusetts 01545.
Bayley
H
H
D000255:Adenosine Triphosphate / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D001048:Aplysia / Q000201:enzymology* / Q000235:genetics; D002384:Catalysis; D007700:Kinetics; D007915:Lepidoptera / Q000235:genetics; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000138:chemical synthesis; D010766:Phosphorylation; D011485:Protein Binding; D011494:Protein Kinases / Q000737:chemistry*; D013379:Substrate Specificity
D009842:Oligopeptides; C037642:kemptide; D000255:Adenosine Triphosphate; D011494:Protein Kinases