Generation of multiple forms of methionyl-tRNA synthetase from the multi-enzyme complex of mammalian aminoacyl-tRNA synthetases by endogenous proteolysis.
Methionyl-tRNA synthetase occurs free and as high-molecular-weight multi-enzyme complexes in rat liver. The free form is purified to near homogeneity by conventional column chromatography and affinity chromatography on tRNA-Sepharose. The native molecular weight of free methionyl-tRNA synthetase is 64 500, based on its sedimentation coefficient of 4.5 S and Stokes radius of 33 A. The free methionyl-tRNA synthetase apparently belongs to alpha-type subunit structure, since the subunit molecular weight is 68 000, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Methionyl-tRNA synthetase is dissociated from the high-molecular-weight synthetase complex by controlled trypsinization, according to Kellermann, O., Viel, C. and Waller, J.P. (Eur. J. Biochem. 88 (1978) 197-204). The dissociated, free methionyl-tRNA synthetase is subsequently purified to near homogeneity. The subunit structure of dissociated methionyl-tRNA synthetase is identical to that of endogenous free methionyl-tRNA synthetase. Anti-serum raised against Mr 104 000 protein in the synthetase complex, specifically inhibited methionyl-tRNA synthetase in both the free and the high-molecular-weight forms to the same extent. These results suggest that the occurrence of multiple forms of methionyl-tRNA synthetases in mammalian cells may, in part, be due to proteolytic cleavage.
D000604:Amino Acyl-tRNA Synthetases / Q000302:isolation & purification*; D000818:Animals; D002499:Centrifugation, Density Gradient; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002845:Chromatography; D004591:Electrophoresis, Polyacrylamide Gel; D007158:Immunologic Techniques; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D008297:Male; D008718:Methionine-tRNA Ligase / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D051381:Rats; D011920:Rats, Inbred WF; D014357:Trypsin / Q000378:metabolism
D046911:Macromolecular Substances; D014357:Trypsin; D000604:Amino Acyl-tRNA Synthetases; D008718:Methionine-tRNA Ligase