Two Enzymically Active Forms of Glycyl‐tRNA Synthetase from Bacillus brevis

Using sucrose density centrifugation and gel filtration of a 105000 × g supernatant of Bacillus brevis two enzymic activities of glycyl‐tRNA synthetase were separated. Enzyme catalyzing the aminoacylation of tRNA (E 1 ) elutes in a high‐molecular‐weight region. Enzyme active in glycylhydroxamate formation (E 2 ) 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 E 1 and E 2 were purified to a nearly homogeneous state. Sedimentation coefficients ( S w.20 ) were found to be 8.6 S and 3.6 S and molecular weights 226000 and 66000 for E 1 and E 2 , respectively. During storage, E 1 dissociates into two components, one of which has electrophoretic mobility identical to E 2 . The molecular weight of the other component is about 160000. Electrophoresis of E 1 in the presence of sodium dodecylsulfate reveals two bands corresponding to molecular weights of 81000 and 30000. Under these conditions, E 2 dissociates into a polypeptide with a molecular weight of 30000. Valine was found to be the N‐terminal amino acid for E 2 and both valine and glutamic acid were N‐terminal amino acids for E 1 . It is concluded that E 1 is a tetrameric protein consisting of two large and two small subunits (α 2 β 2 ). E 2 is a component of E 1 with a structural formula α 2 .