Abstract
Knowledge of the molecular mechanisms of specific bacterial virulence factors can significantly contribute to antibacterial drug discovery. Helicobacter pylori is a Gram-negative microaerophilic bacterium that infects almost half of the world's population, leading to gastric disorders and even gastric cancer. H. pylori expresses a series of virulence factors in the host, among which high-temperature requirement A (HpHtrA) is a newly identified serine protease secreted by H. pylori. HpHtrA cleaves the extracellular domain of the epithelial cell surface adhesion protein E-cadherin and disrupts gastric epithelial cell junctions, allowing H. pylori to access the intercellular space. Here we report the first crystal structure of HpHtrA at 3.0 Å resolution. The structure revealed a new type of HtrA protease trimer stabilized by unique N-terminal domain swapping distinct from other known HtrA homologs. We further observed that truncation of the N terminus completely abrogates HpHtrA trimer formation as well as protease activity. In the presence of unfolded substrate, HpHtrA assembled into cage-like 12-mers or 24-mers. Combining crystallographic, biochemical, and mutagenic data, we propose a mechanistic model of how HpHtrA recognizes and cleaves the well-folded E-cadherin substrate. Our study provides a fundamental basis for the development of anti-H. pylori agents by using a previously uncharacterized HtrA protease as a target.
Highlights
Knowledge of the molecular mechanisms of specific bacterial virulence factors can significantly contribute to antibacterial drug discovery
All mutants showed decreased proteolytic activity against E-cadherin compared with WT HpHtrA, implying that all of these domains might participate in the E-cadherin cleavage process (Fig. 1D)
Recent studies demonstrated that H. pylori utilizes a novel secreted serine protease, HtrA, to cleave the host Occludin, Claudin-8, as well as E-cadherin proteins, which breaks down the E-cadherin– based adherens junctions and tight junctions between gastric epithelial cells to disintegrate the epithelial barrier
Summary
Knowledge of the molecular mechanisms of specific bacterial virulence factors can significantly contribute to antibacterial drug discovery. The strategy seeks to interfere with the bacterial virulence factors that promote infection without threatening their existence. This leads to reduced selective pressure for drug-resistant mutations. Development of antivirulence drugs requires an in-depth understanding of the structures and functions of virulence factors in disease processes. Some virulence factors have been recognized as potential drug targets to eradicate H. pylori infection [8, 9]. High-temperature requirement A protein of H. pylori (HpHtrA) is a newly identified virulence factor that helps H. pylori to efficiently break through the gastric epithelium by cleaving proteins within the epithelial tight junction (occludin and claudin-8) and adherens junction (E-cadherin) [10, 11]. Structural and functional characterization of this virulence factor may facilitate the design of new types of antiH. pylori drugs
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