The molecular structure of the glycoside hydrolase domain of Cwp19 from Clostridium difficile.

William J Bradshaw,K Ravi Acharya,Clifford C Shone,April K Roberts,Jonathan M Kirby

doi:10.1111/febs.14310

William J Bradshaw, K Ravi Acharya + Show 3 more

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https://doi.org/10.1111/febs.14310

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Journal: FEBS Journal	Publication Date: Nov 17, 2017
Citations: 13	License type: CC BY 4.0

Affiliation: University of Bath, Public Health England

Abstract

Clostridium difficile is a burden to healthcare systems around the world, causing tens of thousands of deaths annually. The S‐layer of the bacterium, a layer of protein found of the surface of cells, has received a significant amount of attention over the past two decades as a potential target to combat the growing threat presented by C. difficile infections. The S‐layer contains a wide range of proteins, each of which possesses three cell wall‐binding domains, while many also possess a “functional” region. Here, we present the high resolution structure of the functional region of one such protein, Cwp19 along with preliminary functional characterisation of the predicted glycoside hydrolase. Cwp19 has a TIM barrel fold and appears to possess a high degree of substrate selectivity. The protein also exhibits peptidoglycan hydrolase activity, an order of magnitude slower than that of lysozyme and is the first member of glycoside hydrolase‐like family 10 to be characterised. This research goes some way to understanding the role of Cwp19 in the S‐layer of C. difficile.DatabaseStructural data are available in the PDB under the accession numbers 5OQ2 and 5OQ3.

Highlights

The Gram-positive “superbug” Clostridium difficile has received significant media attention in recent decades as the primary causative agent of antibiotic-associated diarrhoea
The structure of Cwp19-fr has been determined by selenium single-wavelength anomalous diffraction (Se-SAD) and to a high resolution with native data using a construct coding for residues 27-401, electron density is only visible for residues 28-388 across the two structures presented here
The Se-SAD structure of Cwp19-fr has been determined to a resolution of 2.3 A and contains two protein chains in the asymmetric unit with two phosphate ions, a PEG molecule and 136 water molecules, while the high resolution native structure has been determined to 1.35 A with one protein chain in the asymmetric unit, two PEG molecules, a chloride ion and 385 water molecules

Summary

Introduction

The Gram-positive “superbug” Clostridium difficile has received significant media attention in recent decades as the primary causative agent of antibiotic-associated diarrhoea. The bacterium presents a significant burden to healthcare systems, causing tens of thousands of deaths globally each year [4]. This demonstrates that a greater understanding of the bacterium is required for the development of novel strategies to combat C. difficile infections. HMW SLP is responsible for binding to the cell wall and possesses three cell wall-binding domains (pfam 04122, CWB2). There are 28 SlpA paralogues in the C. difficile genome, each of which possesses three CWB2 domains, and many possess “functional” regions [5,10,11].

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