Deletion of the putative effector region of Era, an essential GTP-binding protein in Escherichia coli, causes a dominant-negative phenotype.

Abstract

Era is an essential gene in E. coli, encoding a GTP-binding protein of unknown function. In the present work, a mutant designated Era-dE, for deletion of effector region is described. This is the first and only known era allele that confers a dominant-negative phenotype. Phenotypic analysis of the mutant showed that overproduction of Era-dE caused a dominant inhibition of growth when TCA cycle intermediates such as succinate, pyruvate, malate, alpha-ketoglutarate, and fumarate were provided as the sole carbon source. Examination of the macromolecular composition of cells overexpressing the mutant showed protein, DNA, and ATP levels expected for cells growing at slow rates. The response of cells expressing Era-dE to different stress conditions was studied by examining the rates of synthesis of stress-inducible proteins. Interestingly, when subjected to succinate starvation, cells expressing Era-dE showed a defective carbon starvation response, whereas response to glucose starvation was similar to that seen in control cells. Taken together with previous results, these studies indicate that Era is perhaps involved in multiple cellular processes and Era-dE disrupts more than one of these functions. Furthermore, it appears that some possible functions of Era include regulation of the TCA cycle and response to carbon starvation.