The Escherichia coli GTPase CgtAE cofractionates with the 50S ribosomal subunit and interacts with spot, a ppGpp synthetase/hydrolase

Abstract

CgtAE/ObgE/YhbZ is an Escherichia coli guanine nucleotide binding protein of the Obg/GTP1 subfamily whose members have been implicated in a number of cellular functions including GTP-GDP sensing, sporulation initiation, and translation. Here we describe a kinetic analysis of CgtAE with guanine nucleotides and show that its properties are similar to those of the Caulobacter crescentus homolog CgtAC. CgtAE binds both GTP and GDP with moderate affinity, shows high guanine nucleotide exchange rate constants for both nucleotides, and has a relatively low GTP hydrolysis rate. We show that CgtAE is associated predominantly with the 50S ribosomal subunit. Interestingly, CgtAE copurifies with SpoT, a ribosome-associated ppGpp hydrolase/synthetase involved in the stress response. The interaction between CgtAE and SpoT was confirmed by reciprocal coprecipitation experiments and by two-hybrid assays. These studies raise the possibility that the ribosome-associated CgtA E is involved in the SpoT-mediated stress response.