A temperature-sensitive glycyl-transfer ribonucleic acid synthetase mutant of Escherichia coli

Abstract

A method of selecting temperature-sensitive mutants of Escherichia coli which makes use of a double temperature shift combined with suicide as a result of incorporated 3H-uridine is described. One mutant selected in this way shows restricted growth at 42 °C resulting from early inhibition of protein and ribonucleic acid (RNA) synthesis. In vitro assays of aminoacyl-transfer RNA synthetase activity indicated a complete absence of active glycyl-transfer RNA synthetase in the mutant. This enzyme activity in the mutant was distinguishable from the wild type by its rapid inactivation at 28 °C in cell-free extracts. Infection of the temperature-sensitive mutant with bacteriophage T4 did not alter the heat-sensitivity of the mutant enzyme. The mutation is 72% cotransducible with the xyl locus.