Chemical measurement of steady-state levels of ten aminoacyl-transfer ribonucleic acid synthetases in Escherichia coli

Abstract

Polypeptide chains of 10 aminoacyl-tRNA synthetases (those for arginine, glutamine, glutamic acid, glycine, isoleucine, leucine, lysine, phenylalanine, threonine and valine) were identified in lysates of E. coli resolved by the O''Farrell 2-dimensional gel system. By labeling cells uniformly with [14C]glucose and by measuring the total amounts of these polypeptides by their radioactivity, estimations of the steady-state, molecular amounts of these enzymes were made and compared to the number of ribosomes and elongation factors in these cells. Portions of a reference culture grown on glucose and labeled with [14C]leucine or [35S]sulfate were mixed with 4 cultures grown in widely different media containing [3H]leucine or [3H]leucine plus [3H]isoleucine. From the isotope ratios of the total protein and of the spots containing the synthetase chains, the chemical amount of each synthetase relative to that of the reference culture was determined. The results, where comparable, show reasonable agreement with enzyme activity measurements. In general these synthetases each exhibit a positive correlation with growth rate in unrestricted media, indicating a strong tendency for the levels of tRNA, synthetases, elongation factors and ribosomes to remain approximately, though not exactly, in balance at different growth rates.