Protein kinase C phosphorylates Ser152, Ser156 and Ser163 but not Ser160 of MARCKS in rat brain.
MARCKS is one of the major physiological substrates of PKC and was reported to be phosphorylated by PKC at 4 serine residues that are within the CaM-binding region (Graff et al., J. Biol. Chem. 264, 11912, 1989). Using MARCKS from rat brain and a synthetic peptide of 25 amino acids containing all 4 of the serine residues, we investigate the differences in phosphorylation by PKC isozymes I, II and III. Tryptic peptide analysis of PKC phosphorylated MARCKS or peptide, we found 32P was in peptides of (K)S152FK, (R)FS156FK and LS160GFS163FK. Further digestion of LSGFSFK with alpha-chymotrypsin revealed that 32P incorporation occurred only at Ser163 but not at Ser160. The initial rates and stoichiomatry of phosphorylation of Ser152 and Ser156 were twice as those of Ser163 using either one of the three PKC isozymes. These results indicate that in vitro, PKC phosphorylates MARCKS only at three sites, but not at Ser160 as that reported previously, and there was no preferential phosphorylation of MARCKS by either PKC isozyme I, II or III.
Heemskerk
F M
FM
Endocrinology and Reproduction Research Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892.
Chen
H C
HC
Huang
F L
FL
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D010446:Peptide Fragments / Q000302:isolation & purification; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D012694:Serine; D013379:Substrate Specificity; D014357:Trypsin
D047908:Intracellular Signaling Peptides and Proteins; C000619712:Marcks protein, rat; D008565:Membrane Proteins; D010446:Peptide Fragments; D010455:Peptides; D011506:Proteins; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D012694:Serine; D011493:Protein Kinase C; D014357:Trypsin