Demonstration and purification of a myristoyl-CoA binding protein from bovine cardiac muscle.
Protein myristoylation refers to the co-translational addition of myristoyl group to an amino-terminal glycine residue of a protein by the enzyme myristoyl-CoA:protein N-myristoyltransferase (NMT). The myristoylation reaction depends on the availability of the cellular pools of coenzyme A and myristate and their subsequent formation of myristoyl-CoA, the substrate of NMT. In the present study a myristoyl-CoA binding protein (MCBP) was purified using various column chromatographies: hydroxylapatite, DEAE Sepharose CL-6B and Sephacryl S-300 gel filtration. The purified protein exhibited an apparent molecular mass of 50 kDa on SDS-polyacrylamide gel electrophoresis. Incubation of protein with [1-(14)C]myristoyl-CoA followed by denaturing gel electrophoresis, fluorography and treatment with hydroxylamine yielded results that are highly suggestive of a covalent ester-linked acyl-protein complex. This complex formation was not observed in the crude cytosolic fractions. The addition of cytosolic fraction to a progressing acyl-protein complex, resulted in deacylation suggesting a role for thioesterase or/proteinases in the regulation of the acylation reaction in bovine cardiac muscle. The acyl-protein complex formation was not inhibited by NIP71, a 71 kDa NMT inhibitory protein from bovine brain.
Raju
R V
RV
Department of Pathology, College of Medicine, University of Saskatchewan, Royal University Hospital, Saskatoon, Canada.
Sharma
R K
RK
D000214:Acyl Coenzyme A / Q000378:metabolism*; D000217:Acyltransferases / Q000037:antagonists & inhibitors; D000818:Animals; D002417:Cattle; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008970:Molecular Weight; D009206:Myocardium / Q000378:metabolism*; D011485:Protein Binding; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*
D000214:Acyl Coenzyme A; D011506:Proteins; C040349:S-tetradecanoyl-coenzyme A; D000217:Acyltransferases; C050480:glycylpeptide N-tetradecanoyltransferase