Two enzymically active forms of glycyl-tRNA synthetase from Bacillus brevis. Purification and properties.
Using sucrose density centrifugation and gel filtration of a 105000 X g supernatant of Bacillus brevis two enzymic activities of glycyl-tRNA synthetase were separated. Enzyme catalyzing the aminoacylation of tRNA (E1) elutes in a high-molecular-weight region. Enzyme active in glycylhydroxamate formation (E2) elutes from a Sephadex gel column and sediments in sucrose density gradient in a region of relatively low molecular weight. The presence of two enzymic activities does not depend on the method of cell disruption; their proportion does not change when protease inhibitor (diisopropylphosphorofluoridate) is added to the extraction buffer. Both E1 and E2 were purified to a nearly homogeneous state. Sedimentation coefficients (sw,20) were found to be 8.6 S and 3.6 S and molecular weights 226000 and 66000 for E1 and E2, respectively. During storage, E1 dissociates into two components, one of which has electrophoretic mobility identical to E2. The molecular weight of the other component is about 1600000. Electrophoresis of E1 in the presence of sodium dodecylsulfate reveals two bands corresponding to molecular weights of 81000 and 30000. Under these conditions, E2 dissociates into a polypeptide with a molecular weight of 30000. Valine was found to be the N-terminal amino acid for E2 and both valine and glutamic acid were N-terminal amino acids for E1. It is concluded that E1 is a tetrameric protein consisting of two large and two small subunits (alpha2beta2). E2 is a component of E1 with a structural formula alpha2.
D000596:Amino Acids / Q000032:analysis; D000604:Amino Acyl-tRNA Synthetases; D001407:Bacillus / Q000201:enzymology* / Q000648:ultrastructure; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D003600:Cytosol / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D005971:Glutamates / Q000032:analysis; D006032:Glycine-tRNA Ligase / Q000032:analysis / Q000302:isolation & purification; D007531:Isoflurophate; D046911:Macromolecular Substances; D008970:Molecular Weight; D012967:Sodium Dodecyl Sulfate; D014633:Valine / Q000032:analysis
D000596:Amino Acids; D005971:Glutamates; D046911:Macromolecular Substances; D007531:Isoflurophate; D012967:Sodium Dodecyl Sulfate; D000604:Amino Acyl-tRNA Synthetases; D006032:Glycine-tRNA Ligase; D014633:Valine