Dr Stéphane Mesnage
School of Biosciences
Senior Lecturer
+44 114 222 4405
Full contact details
School of Biosciences
Firth Court
Western Bank
Sheffield
S10 2TN
- Profile
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- 2019 - present: Senior Lecturer, School of Biosciences, University of Sheffield, UK
- 2012 - 2018: Lecturer, Dept. of Molecular Biology and Biotechnology, University of Sheffield, UK
- 2010 - 2012: Marie Curie (IEF) Fellow, Dept. of Molecular Biology and Biotechnology, University of Sheffield, UK
- 2002 - 2010: INSERM Chargé de Recherches, University of Paris 6, France
- 2000 - 2002: EMBO Research assistant, John Innes Centre, Norwich, UK
- 1996 - 2000: PhD, Pasteur Institute, Paris, France
- Research interests
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My work is focused on the study of the bacterial cell wall using essentially two Gram-positive pathogens (Staphylococcus aureus, Enterococcus faecalis) as models. I am interested in three major areas of research.
Analysis of protein-cell wall interactions
The essential cell wall peptidoglycan polymer network forms a scaffold for the display of a large variety of proteins involved in cell division, metabolism, cell-cell communication and in pathogen interaction with the host. Although several multimodular cell wall binding domains have been identified to date in bacteria, both their structural organisation and binding mechanism (binding motif, subcellular localisation) have remained elusive.
The aim of this project is to understand the molecular mechanisms underpinning protein-bacterial cell wall interactions and subcellular localisation of surface proteins.
Beyond these fundamental questions, the aim is to develop a translational research program to (i) develop peptidoglycan binding domains as new functional probes to provide a nanoscale topological map of the cell surface; (ii) provide an experimental rationale to engineer and target therapeutic molecules against pathogens.
This project involves multidisciplinary approaches across several disciplines including molecular biology, biochemistry, biophysics, structural biology and state-of-the-art super resolution microscopy.
Control of peptidoglycan hydrolysis during growth
During growth, the insertion of new precursors and separation of daughter cells requires limited cleavage of the peptidoglycan molecule by peptidoglycan hydrolases.
Depending on the bond they cleave, these enzymes are classified as N-acetylmuramidases, N-acetylglucosaminidases, N-acetylmuramoyl L-alanine amidases, endopeptidases or carboxypeptidases (Fig. 2). In addition to their contribution to cell growth and division, some peptidoglycan hydrolases play a role in adhesion and in amplification of the inflammatory response by releasing muramyl-peptides.
The lytic activity of some peptidoglycan hydrolases (called autolysins) is potentially lethal and can lead to cell lysis and death. How bacteria control autolytic activities is poorly understood.
I am studying several mechanisms that may be responsible for the control of autolytic activities: (i) the subcellular targeting of autolysins; (ii) the impact of peptidoglycan structural properties on autolytic activities and (iii) the contribution of the electrochemical gradient across the cytoplasmic membrane to the control of peptidoglycan hydrolysis.
We routinely analyse peptidoglycan hydrolase substrate specificity by a combination of rp-HPLC and mass spectrometry. The enzymatic activity of autolysins is measured spectrophotometrically.
Role of Enterococcus faecalis cell wall metabolism in the interaction with the host
Lysozyme is the major component of the innate immune system and represents the first line of defense against pathogens. E. faecalis is highly resistant to lysozyme, with a minimal inhibitory concentration (MIC) above 50 mg/ml.
This extreme resistance has been partly explained by an additive effect of peptidoglycan O-acetylation and de-N-acetylation, as well as teichoic acid D-alanylation. It was also shown that full lysozyme resistance requires a signalling cascade involving the extracytoplasm
ic function (ECF) sigma factor SigV. Interestingly, E. faecalis strains harbouring multiple deletions in these pathways are still resistant to lysozyme. The identification of novel factors involved in lysozyme resistance is currently underway using random transposon mutagenesis. The mutants isolated will be further characterised in vivo.
- Publications
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Show: Featured publications All publications
Featured publications
Journal articles
- Bacterial size matters: multiple mechanisms controlling septum cleavage and diplococcus formation are critical for the virulence of the opportunistic pathogen Enterococcus faecalis. PLoS Pathogens, 13(7). View this article in WRRO
- Molecular basis for bacterial peptidoglycan recognition by LysM domains.. Nat Commun, 5, 4269. View this article in WRRO
- Zebrafish as a novel vertebrate model to dissect enterococcal pathogenesis.. Infect Immun, 81(11), 4271-4279. View this article in WRRO
- Super-resolution microscopy reveals cell wall dynamics and peptidoglycan architecture in ovococcal bacteria. Molecular Microbiology, 82(5), 1096-1109.
- Cleavage specificity of Enterococcus faecalis EnpA (EF1473), a peptidoglycan endopeptidase related to the LytM/lysostaphin family of metallopeptidases.. J Mol Biol, 398(4), 507-517.
- Impact of peptidoglycan O-acetylation on autolytic activities of the Enterococcus faecalis N-acetylglucosaminidase AtlA and N-acetylmuramidase AtlB.. FEBS Lett, 583(18), 3033-3038.
- Role of N-acetylglucosaminidase and N-acetylmuramidase activities in Enterococcus faecalis peptidoglycan metabolism.. J Biol Chem, 283(28), 19845-19853.
All publications
Journal articles
- Dissecting the Enterococcal Polysaccharide Antigen (EPA) structure to explore innate immune evasion and phage specificity. Carbohydrate Polymers, 347, 122686-122686.
- A diverse set of Enterococcus-infecting phage provides insight into phage host-range determinants. Virus Research, 347, 199426-199426.
- Clostridioides difficile canonical L,D-transpeptidases catalyse a novel type of peptidoglycan cross-links and are not required for beta-lactam resistance. Journal of Biological Chemistry, 105529-105529.
- Unusual 1-3 peptidoglycan cross-links in Acetobacteraceae are made by L,D-transpeptidases with a catalytic domain distantly related to YkuD domains. Journal of Biological Chemistry, 105494-105494.
- A moonlighting role for LysM peptidoglycan binding domains underpins Enterococcus faecalis daughter cell separation. Communications Biology, 6(1).
- Human serum triggers antibiotic tolerance in Staphylococcus aureus. Nature Communications, 13(1).
- Molecular basis for substrate recognition and septum cleavage by AtlA, the major N-acetylglucosaminidase of Enterococcus faecalis. Journal of Biological Chemistry.
- Structural characterization of EnpA D,L-Endopeptidase from Enterococcus faecalis prophage provides insights into substrate specificity of M23 peptidases. International Journal of Molecular Sciences, 22(13).
- A family of T6SS antibacterial effectors related to L,D-transpeptidases targets the peptidoglycan. Cell Reports, 31(12). View this article in WRRO
- Complete Structure of the Enterococcal Polysaccharide Antigen (EPA) of Vancomycin-Resistant Enterococcus faecalis V583 Reveals that EPA Decorations Are Teichoic Acids Covalently Linked to a Rhamnopolysaccharide Backbone. mBio, 11(2). View this article in WRRO
- Two-site recognition of Staphylococcus aureus peptidoglycan by lysostaphin SH3b. Nature Chemical Biology. View this article in WRRO
- Peptidoglycan Production by an Insect-Bacterial Mosaic. Cell, 179(3), 703-712.e7.
- Identification of novel bacteriophages with therapeutic potential targeting Enterococcus faecalis. Infection and Immunity. View this article in WRRO
- Decoration of the enterococcal polysaccharide antigen EPA is essential for virulence, cell surface charge and interaction with effectors of the innate immune system. PLOS Pathogens, 15(5). View this article in WRRO
- Molecular imaging of glycan chains couples cell-wall polysaccharide architecture to bacterial cell morphology. Nature Communications, 9(1). View this article in WRRO
- Molecular coordination of staphylococcus aureus cell division. eLife, 7. View this article in WRRO
- RNA-seq and Tn-seq reveal fitness determinants of vancomycin-resistant Enterococcus faecium during growth in human serum. BMC Genomics, 18(1). View this article in WRRO
- Bacterial size matters: multiple mechanisms controlling septum cleavage and diplococcus formation are critical for the virulence of the opportunistic pathogen Enterococcus faecalis. PLoS Pathogens, 13(7). View this article in WRRO
- The mechanism behind the selection of two different cleavage sites in NAG-NAM polymers.. IUCrJ, 4(Pt 2), 185-198. View this article in WRRO
- Towards an automated analysis of bacterial peptidoglycan structure. Analytical and Bioanalytical Chemistry, 409(2), 551-560. View this article in WRRO
- DUF3380 domain from a Salmonella phage endolysin shows potent N -acetylmuramidase activity. Applied and Environmental Microbiology, 82(16), 4975-4981. View this article in WRRO
- Structural and Enzymatic Characterization of ABgp46, a Novel Phage Endolysin with Broad Anti-Gram-Negative Bacterial Activity. Frontiers in Microbiology, 7. View this article in WRRO
- Bacterial Cell Enlargement Requires Control of Cell Wall Stiffness Mediated by Peptidoglycan Hydrolases. mBio, 6(4). View this article in WRRO
- Clostridium difficile surface proteins are anchored to the cell wall using CWB2 motifs that recognise the anionic polymer PSII. Molecular Microbiology, 96(3), 596-608. View this article in WRRO
- The Aeromonas caviae AHA0618 gene modulates cell length and influences swimming and swarming motility. MicrobiologyOpen, 4(2), 220-234. View this article in WRRO
- Prevalence and Gene Characteristics of Antibodies with Cofactor-induced HIV-1 Specificity. Journal of Biological Chemistry, 290(8), 5203-5213. View this article in WRRO
- Molecular basis for bacterial peptidoglycan recognition by LysM domains.. Nat Commun, 5, 4269. View this article in WRRO
- A cryptic polyreactive antibody recognizes distinct clades of HIV-1 glycoprotein 120 by an identical binding mechanism.. J Biol Chem, 289(25), 17767-17779.
- Zebrafish as a novel vertebrate model to dissect enterococcal pathogenesis.. Infect Immun, 81(11), 4271-4279. View this article in WRRO
- De-O-acetylation of peptidoglycan regulates glycan chain extension and affects in vivo survival of Neisseria meningitidis.. Mol Microbiol, 87(5), 1100-1112.
- N-Acetylmuramoyl-L-alanine Amidase, 1, 1401-1407.
- The lysozyme-induced peptidoglycan N-acetylglucosamine deacetylase PgdA (EF1843) is required for Enterococcus faecalis virulence.. J Bacteriol, 194(22), 6066-6073.
- Super-resolution microscopy reveals cell wall dynamics and peptidoglycan architecture in ovococcal bacteria. Molecular Microbiology, 82(5), 1096-1109.
- The in vitro contribution of autolysins to bacterial killing elicited by amoxicillin increases with inoculum size in Enterococcus faecalis.. Antimicrob Agents Chemother, 55(2), 910-912.
- Cleavage specificity of Enterococcus faecalis EnpA (EF1473), a peptidoglycan endopeptidase related to the LytM/lysostaphin family of metallopeptidases.. J Mol Biol, 398(4), 507-517.
- Impact of peptidoglycan O-acetylation on autolytic activities of the Enterococcus faecalis N-acetylglucosaminidase AtlA and N-acetylmuramidase AtlB.. FEBS Lett, 583(18), 3033-3038.
- Contribution of the autolysin AtlA to the bactericidal activity of amoxicillin against Enterococcus faecalis JH2-2.. Antimicrob Agents Chemother, 53(4), 1667-1669.
- Role of N-acetylglucosaminidase and N-acetylmuramidase activities in Enterococcus faecalis peptidoglycan metabolism.. J Biol Chem, 283(28), 19845-19853.
- Idiosyncratic features in tRNAs participating in bacterial cell wall synthesis.. Nucleic Acids Res, 35(20), 6870-6883. View this article in WRRO
- The Enterococcus hirae Mur-2 enzyme displays N-acetylglucosaminidase activity.. FEBS Lett, 581(4), 693-696.
- Functional analysis of AtlA, the major N-acetylglucosaminidase of Enterococcus faecalis.. J Bacteriol, 188(24), 8513-8519.
- LHP1, the Arabidopsis homologue of HETEROCHROMATIN PROTEIN1, is required for epigenetic silencing of FLC.. Proc Natl Acad Sci U S A, 103(13), 5012-5017.
- Structure-based site-directed mutagenesis of the UDP-MurNAc-pentapeptide-binding cavity of the FemX alanyl transferase from Weissella viridescens.. J Bacteriol, 187(11), 3833-3838.
- In vivo Bacillus anthracis gene expression requires PagR as an intermediate effector of the AtxA signalling cascade.. Int J Med Microbiol, 293(7-8), 619-624.
- Structural analysis and evidence for dynamic emergence of Bacillus anthracis S-layer networks.. J Bacteriol, 184(23), 6448-6456.
- Multiple roles of Arabidopsis VRN1 in vernalization and flowering time control.. Science, 297(5579), 243-246.
- Developmental switch of S-layer protein synthesis in Bacillus anthracis.. Mol Microbiol, 43(6), 1615-1627.
- Bacillus anthracis cell envelope components.. Curr Top Microbiol Immunol, 271, 87-113.
- Plasmid-encoded autolysin in Bacillus anthracis: modular structure and catalytic properties.. J Bacteriol, 184(1), 331-334.
- A general strategy for identification of S-layer genes in the Bacillus cereus group: molecular characterization of such a gene in Bacillus thuringiensis subsp. galleriae NRRL 4045.. Microbiology, 147(Pt 5), 1343-1351.
- Conserved anchoring mechanisms between crystalline cell surface S-layer proteins and secondary cell wall polymers in Gram-positive bacteria? Response. TRENDS MICROBIOL, 9(2), 49-50.
- Bacterial SLH domain proteins are non-covalently anchored to the cell surface via a conserved mechanism involving wall polysaccharide pyruvylation.. EMBO J, 19(17), 4473-4484.
- Cell surface-exposed tetanus toxin fragment C produced by recombinant Bacillus anthracis protects against tetanus toxin.. Infect Immun, 67(9), 4847-4850.
- Bacillus anthracis surface: capsule and S-layer.. J Appl Microbiol, 87(2), 251-255.
- The S-layer homology domain as a means for anchoring heterologous proteins on the cell surface of Bacillus anthracis.. J Appl Microbiol, 87(2), 256-260.
- Production and cell surface anchoring of functional fusions between the SLH motifs of the Bacillus anthracis S-layer proteins and the Bacillus subtilis levansucrase.. Mol Microbiol, 31(3), 927-936.
- The capsule and S-layer: two independent and yet compatible macromolecular structures in Bacillus anthracis.. J Bacteriol, 180(1), 52-58.
- IV. Molecular biology of S-layers. FEMS Microbiology Reviews, 20(1-2), 47-98.
- Molecular biology of S-layers.. FEMS Microbiol Rev, 20(1-2), 47-98.
- Molecular characterization of the Bacillus anthracis main S-layer component: evidence that it is the major cell-associated antigen.. Mol Microbiol, 23(6), 1147-1155.
- Nucleotide sequences of four pathogen-induced alfalfa peroxidase-encoding cDNAs.. Gene, 170(2), 213-216.
- Exploring the role of E. faecalis enterococcal polysaccharide antigen (
EPA ) and lipoproteins in evasion of phagocytosis. Molecular Microbiology.
- Structural and Biochemical Characterization of a New Phage-Encoded Muramidase, KTN6 Gp46. PHAGE.
- 1H, 13C, and 15N resonance assignments of a conserved putative cell wall binding domain from Enterococcus faecalis. Biomolecular NMR Assignments.
- β-Barrel proteins tether the outer membrane in many Gram-negative bacteria. Nature Microbiology.
- PGFinder, a novel analysis pipeline for the consistent, reproducible, and high-resolution structural analysis of bacterial peptidoglycans. eLife, 10.
Chapters
- PGFinder, an Open-Source Software for Peptidoglycomics: The Structural Analysis of Bacterial Peptidoglycan by LC-MS, Protein Bioinformatics (pp. 111-132). Springer US
- N-Acetylmuramoyl-l-alanine Amidase, Handbook of Proteolytic Enzymes (pp. 1401-1407). Elsevier
Conference proceedings papers
- Determining the structure of the enterococcal polysaccharide antigen. FEBS OPEN BIO, Vol. 12 (pp 41-41)
- A Family of T6SS Antibacterial Effectors related to L,D-transpeptidases Targets the Peptidoglycan. BIOPHYSICAL REVIEWS, Vol. 13(6) (pp 1408-1409)
- Automated Analysis of Bacterial Peptidoglycan Structure. GLYCOBIOLOGY, Vol. 26(12) (pp 1457-1457)
Preprints
- The Enterococcal Polysaccharide Antigen: from structure to biosynthesis and function, Cold Spring Harbor Laboratory.
- Exploring the role ofE. faecalisEnterococcal Polysaccharide Antigen (EPA) and lipoproteins in evasion of phagocytosis, Cold Spring Harbor Laboratory.
- Exploring the role of E. faecalis Enterococcal Polysaccharide Antigen (EPA) and lipoproteins in evasion of phagocytosis, Authorea, Inc..
- Unusual 1-3 peptidoglycan cross-links inAcetobacteraceaeare made by L,D-transpeptidases with a catalytic domain distantly related to YkuD domains, Cold Spring Harbor Laboratory.
- Human serum triggers antibiotic tolerance in Staphylococcus aureus, Cold Spring Harbor Laboratory.
- PGfinder, a novel analysis pipeline for the consistent, reproducible and high-resolution structural analysis of bacterial peptidoglycans.
- A Superfamily of T6SS Antibacterial Effectors Displaying L,D-carboxypeptidase Activity Towards Peptidoglycan, Cold Spring Harbor Laboratory.
- A Superfamily of T6SS Antibacterial Effectors Displaying L,D-Carboxypeptidase Activity Towards Peptidoglycan.
- Decoration of the enterococcal polysaccharide antigen EPA is essential for virulence, cell surface charge and resistance to innate immunity, Cold Spring Harbor Laboratory.
- Fitness determinants of vancomycin-resistantEnterococcus faeciumduring growth in human serum, Cold Spring Harbor Laboratory.
- Bacterial size matters: multiple mechanisms controlling septum cleavage and diplococcus formation are critical for the virulence of the opportunistic pathogen Enterococcus faecalis. PLoS Pathogens, 13(7). View this article in WRRO