A cardiolipin-activated protein kinase from rat liver structurally distinct from the protein kinases C.
A cardiolipin- and protease-activated protein kinase (PAK) has been isolated from cytoplasmic extracts of rat liver. The enzyme (PAK-1) phosphorylates the ribosomal protein S6-(229-239) peptide analogue and can be activated by limited proteolysis. Partial amino acid sequences of tryptic peptides derived from both the purified 116-kDa PAK-1 holoenzyme and its active catalytic fragment reveal that the catalytic domain is most related (50-58% identity) to the protein kinase C family. PAK-1 has protein and peptide substrate specificities distinct from those of known protein kinase C isoforms and is insensitive to inhibition by the protein kinase C-alpha-(19-31) pseudosubstrate peptide. Phosphatidylserine, diacylglycerol, and phorbol ester do not activate PAK-1 toward the S6 peptide substrate. However, other acidic phospholipids, the most effective being cardiolipin, activate PAK-1 to a similar extent as trypsin. The PAK-1 catalytic activities generated through activation by cardiolipin or limited proteolysis were kinetically similar, with Km values of 3.6 and 3.4 microM, respectively, for the S6-(229-239) peptide substrate. However, differences were observed in the catalytic activities with protamine sulfate and the glycogen synthase-(1-12) peptide analogue as substrates. It was concluded that PAK-1 is a phospholipid-regulated protein kinase with a primary structure, substrate specificity, and mechanism of regulation in vitro distinct from those of any known member of the protein kinase C superfamily.
Morrice
N A
NA
Russell Grimwade School of Biochemistry, University of Melbourne, Parkville, Victoria, Australia.
Gabrielli
B
B
Kemp
B E
BE
Wettenhall
R E
RE
D000595:Amino Acid Sequence; D000818:Animals; D002308:Cardiolipins / Q000494:pharmacology*; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D008099:Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010449:Peptide Mapping; D010718:Phosphatidylserines / Q000378:metabolism; D011493:Protein Kinase C / Q000737:chemistry / Q000378:metabolism*; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D051381:Rats; D011915:Rats, Inbred BUF; D013379:Substrate Specificity; D014357:Trypsin
D002308:Cardiolipins; D010446:Peptide Fragments; D010718:Phosphatidylserines; D011494:Protein Kinases; D011493:Protein Kinase C; D014357:Trypsin