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

J Comput Aided Mol Des. 2024 Nov 25;39(1):1. doi: 10.1007/s10822-024-00577-x.

Abstract

This study aimed to reveal interactions of the stress response sigma subunits (factors) σD and σH 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 σH subunit led to the creation of a panel of σH variants. Their ability to initiate transcription from naturally occurring hybrid σD/σH-dependent promoter Pcg0441 and two control canonical promoters (σD-dependent PrsdA and σH-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 PD35H10 combining the -10 element of the PuvrD3 promoter and the -35 element of the PrsdA promoter. This artificial hybrid promoter PD35-rsdAH10-uvrD3 showed almost optimal properties needed for the bio-orthogonal transcription (not interfering with the native biological processes).

Keywords: Bio-orthogonal transcription; Corynebacterium; Promoter; Sigma factor.

MeSH terms

  • Bacterial Proteins* / chemistry
  • Bacterial Proteins* / genetics
  • Bacterial Proteins* / metabolism
  • Computer Simulation
  • Corynebacterium glutamicum* / genetics
  • DNA-Directed RNA Polymerases / chemistry
  • DNA-Directed RNA Polymerases / genetics
  • DNA-Directed RNA Polymerases / metabolism
  • Gene Expression Regulation, Bacterial
  • Molecular Dynamics Simulation*
  • Point Mutation*
  • Promoter Regions, Genetic*
  • Sigma Factor* / chemistry
  • Sigma Factor* / genetics
  • Sigma Factor* / metabolism
  • Stress, Physiological / genetics
  • Transcription, Genetic

Substances

  • Sigma Factor
  • Bacterial Proteins
  • DNA-Directed RNA Polymerases