An in vivo effect of the metabolites L-alanine and glycyl-L-leucine on the properties of the lysyl-tRNA synthetase from Escherichia coli K-12. II. Kinetic evidence

Abstract

Wild-type Escherichia coli K-12 was grown in minimal medium alone or with the addition of 20 mM L-alanine or 3 mM glycyl-L-leucine. A lysyl-tRNA synthetase mutant strain was grown in minimal medium containing 20mM L-alanine. The lysyl-tRNA synthetase from these strains was purified to 70-90% of homogeneity. Kinetic studies comparing the effect of thermal and urea inactivation on these different lysyl-tRNA synthetase preparations and measurement of the Michaelis constant for lysine and transfer RNA indicated that growth of Escherichia coli in the presence of alanine and glycyl-L-leucine induces an alteration in the properties of the synthetase. Measurement of the apparent Km for ATP at pH 7.25 indicates lysyl-tRNA synthetase has two two binding sites for this substrate, and further studies indicated a dependence of the apparent Km for lysine on the ATP concentration.