Promoter recognition specificity of Corynebacterium glutamicum stress response sigma factors σ and σ deciphered using computer modeling and point mutagenesis.

This study aimed to reveal interactions of the stress response sigma subunits (factors) σ and σ of RNA polymerase and promoters in Gram-positive bacterium Corynebacterium glutamicum by combining wet-lab obtained data and in silico modeling. Computer modeling-guided point mutagenesis of C. glutamicum σ subunit led to the creation of a panel of σ variants. Their ability to initiate transcription from naturally occurring hybrid σ/σ-dependent promoter Pcg0441 and two control canonical promoters (σ-dependent PrsdA and σ-dependent PuvrD3) was measured and interpreted using molecular dynamics simulations of homology models of all complexes. The results led us to design the artificial hybrid promoter PDH combining the -10 element of the PuvrD3 promoter and the -35 element of the PrsdA promoter. This artificial hybrid promoter PDH showed almost optimal properties needed for the bio-orthogonal transcription (not interfering with the native biological processes).