Definition of the Binding Architecture to a Target Promoter of HP1043, the Essential Master Regulator of Helicobacter pylori.

Annamaria Zannoni,Vincenzo Scarlato,Federica Chiappori,Simone Pelliciari,Francesco Musiani,Davide Roncarati

doi:10.3390/ijms22157848

Abstract

HP1043 is an essential orphan response regulator of Helicobacter pylori orchestrating multiple crucial cellular processes. Classified as a member of the OmpR/PhoB family of two-component systems, HP1043 exhibits a highly degenerate receiver domain and evolved to function independently of phosphorylation. Here, we investigated the HP1043 binding mode to a target sequence in the hp1227 promoter (Php1227). Scanning mutagenesis of HP1043 DNA-binding domain and consensus sequence led to the identification of residues relevant for the interaction of the protein with a target DNA. These determinants were used as restraints to guide a data-driven protein-DNA docking. Results suggested that, differently from most other response regulators of the same family, HP1043 binds in a head-to-head conformation to the Php1227 target promoter. HP1043 interacts with DNA largely through charged residues and contacts with both major and minor grooves of the DNA are required for a stable binding. Computational alanine scanning on molecular dynamics trajectory was performed to corroborate our findings. Additionally, in vitro transcription assays confirmed that HP1043 positively stimulates the activity of RNA polymerase.

Highlights

Helicobacter pylori is a widespread human pathogen recognized as a class I carcinogen by the World Health Organization
The ability of this bacterium to colonize the harsh niche of the stomach and to establish a persistent infection depends on the coordinated expression of several genes, including virulence factors
The alignment of experimentally validated HP1043 binding sites led to the definition of a consensus sequence composed by the repetition of two TTTAAG hexamers separated by a 5 bp spacer, with a lower degree of sequence conservation for the second hemi-site [13]

Summary

Introduction

Helicobacter pylori is a widespread human pathogen recognized as a class I carcinogen by the World Health Organization. It represents the primary cause of severe gastrointestinal diseases such as peptic ulcer, gastric adenocarcinoma, and MALT lymphoma [1,2]. The ability of this bacterium to colonize the harsh niche of the stomach and to establish a persistent infection depends on the coordinated expression of several genes, including virulence factors. Only 17 transcriptional regulators out of approximately 1500 predicted open reading frames (ORFs) have been identified and partially characterized [3]

Methods

Results

Conclusion