DevS, a heme-containing two-component oxygen sensor of Mycobacterium tuberculosis

Area of Science:

• Microbiology
• Biochemistry
• Molecular Biology

Background:

• Mycobacterium tuberculosis exhibits a latent, quiescent state, challenging drug eradication and necessitating prolonged treatment.
• The DevS-DevR two-component system is implicated in mediating cellular responses to anaerobiosis, a condition that can induce bacterial latency.
• Targeting latent tuberculosis requires understanding the molecular mechanisms governing bacterial adaptation and survival.

Purpose of the Study:

• To clone, express, and characterize a truncated DevS protein (DevS642) containing the GAF-A sensor domain and the full-length DevS protein.
• To investigate the heme-binding properties and ligand interactions of DevS, particularly its response to oxygen, nitric oxide, and carbon monoxide.
• To elucidate the role of specific amino acid residues, such as His149, in heme ligation and the influence of domain interactions on DevS function.

Main Methods:

• Cloning and expression of truncated (DevS642) and full-length DevS proteins.
• Heme-binding assays to determine stoichiometry and ligand interactions (O2, NO, CO).
• UV-vis and resonance Raman spectroscopy to analyze wild-type and mutant (H149A) protein structures and heme coordination.

Main Results:

• DevS642 quantitatively binds heme in a 1:1 stoichiometry.
• The ferrous heme-DevS complex reversibly binds O2, NO, and CO.
• Spectroscopic analysis confirmed His149 as the proximal heme ligand, with domain interactions in full-length DevS influencing the distal heme pocket environment.

Conclusions:

• DevS protein binds heme and reversibly interacts with small gas molecules, suggesting a role in sensing environmental changes.
• His149 is crucial for heme ligation, and inter-domain communication within DevS modulates its heme-binding pocket.
• These findings provide insights into the molecular mechanisms of DevS function, potentially aiding in the development of new strategies against latent tuberculosis.