Expression, abundance, and RNA polymerase binding properties of the δ factor of Bacillus subtilis

Abstract

The δ protein is a dispensable subunit of Bacillus subtilis RNA polymerase (RNAP) that has major effects on the biochemical properties of the purified enzyme. In the presence of δ, RNAP displays an increased specificity of transcription, a decreased affinity for nucleic acids, and an increased efficiency of RNA synthesis because of enhanced recycling. Despite these profound effects, a strain containing a deletion of the δ gene (rpoE) is viable and shows no major alterations in gene expression. Quantitative immunoblotting experiments demonstrate that δ is present in molar excess relative to RNAP in both vegetative cells and spores. Expression of rpoE initiates from a single, σ(A)-dependent promoter and is maximal in transition phase. A rpoE mutant strain has an altered morphology and is delayed in the exit from stationary phase. For biochemical analyses we have created derivatives of δ and σ(A) that can be radiolabeled with protein kinase A. Using electrophoretic mobility shift assays, we demonstrate that δ binds core RNAP with an apparent affinity of 2.5 x 106 M-1, but we are unable to demonstrate the formation of a ternary complex containing core enzyme, δ, and σ(A).