Competition among seven Escherichia coli sigma subunits: relative binding affinities to the core RNA polymerase

Abstract

Seven different species of the RNA polymerase σ subunit exist in Escherichia coli, each binding to a single species of the core enzyme and thereby directing transcription of a specific set of genes. To test the σ competition model in the global regulation of gene transcription, all seven E.coli σ subunits have been purified and compared for their binding affinities to the same core RNA polymerase (E). In the presence of a fixed amount of σ⁷⁰, the principal σ for growth-related genes, the level of Eσ⁷⁰ holoenzyme formation increased linearly with the increase in core enzyme level, giving an apparent K_d for the core enzyme of 0.26 nM. Mixed reconstitution experiments in the presence of a fixed amount of core enzyme and increasing amounts of an equimolar mixture of all seven σ subunits indicated that σ⁷⁰ is strongest in terms of core enzyme binding, followed by σ^N, σ^F, σ^E/σ^FecI, σ^H and σ^S in decreasing order. The orders of core binding affinity between σ⁷⁰ and σ^N and between σ⁷⁰ and σ^H were confirmed by measuring the replacement of one core-associated σ by another σ subunit. Taken together with the intracellular σ levels, we tried to estimate the number of each holoenzyme form in growing E.coli cells.