NAPCLI: Non-conventional approaches for peptidoglycan cross-linking inhibition
Lead Research Organisation:
Newcastle University
Department Name: Inst for Cell and Molecular Biosciences
Abstract
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Technical Summary
Peptidoglycan (PG) is an attractive and validated target for antibacterial drug development for two
main reasons. First, it is an essential and unique bacterial cell wall polymer with no counterpart in
human cells, minimizing the risk of drug toxicity. Second, the essential PG synthases are exposed at
the outer surface of the cytoplasmic membrane, making them highly accessible for antibiotic
inhibition. Formation of the PG network requires glycosyltransferases for glycan chain elongation
and transpeptidases for peptide cross-linking. Transpeptidation involves two stem peptides that act
as acyl donor and acceptor substrates, respectively. The acyl donor site is targeted by the
ß-lactams, which form covalent adducts, and this interaction is well characterized. In contrast,
nothing is known on the interaction of the transpeptidases with the acceptor substrate. To combat
the erosion of the activity of ß-lactams, we propose to identify additional drugable sites in the
transpeptidases, including the acceptor binding site, and develop lead antibacterial agents acting
on these sites. Our first objective is to characterize the mode of recognition of the acyl acceptor by
transpeptidases and identify compounds blocking the binding of this substrate. We will use NMR
spectroscopy to map the acceptor site and develop specific inhibitors based on modeling and
virtual screening. Our second objective is to identify the partners of transpeptidases that regulate
the coordinated elongation of glycan chains and cross-linking of stem peptides. This will allow us to
select additional drugable sites in transpeptidases and associated proteins within the PG
polymerization complexes. We will map key interactions by FRET analyses in live bacteria producing
fluorescent proteins and by in vitro transpeptidase/glycosyltransferase assays in complexes
obtained by tandem-affinity purification. Microfluidic cultures and time-lapse microscopy will assess
the impact of inhibitors on cell division and viability. The interaction of lead compounds with their
targets will be characterized by X-ray crystallography. These complementary approaches will
enable the consortium to develop novel strategies for transpeptidase inhibition and obtain leads
active against ß-lactam-resistant bacteria.
main reasons. First, it is an essential and unique bacterial cell wall polymer with no counterpart in
human cells, minimizing the risk of drug toxicity. Second, the essential PG synthases are exposed at
the outer surface of the cytoplasmic membrane, making them highly accessible for antibiotic
inhibition. Formation of the PG network requires glycosyltransferases for glycan chain elongation
and transpeptidases for peptide cross-linking. Transpeptidation involves two stem peptides that act
as acyl donor and acceptor substrates, respectively. The acyl donor site is targeted by the
ß-lactams, which form covalent adducts, and this interaction is well characterized. In contrast,
nothing is known on the interaction of the transpeptidases with the acceptor substrate. To combat
the erosion of the activity of ß-lactams, we propose to identify additional drugable sites in the
transpeptidases, including the acceptor binding site, and develop lead antibacterial agents acting
on these sites. Our first objective is to characterize the mode of recognition of the acyl acceptor by
transpeptidases and identify compounds blocking the binding of this substrate. We will use NMR
spectroscopy to map the acceptor site and develop specific inhibitors based on modeling and
virtual screening. Our second objective is to identify the partners of transpeptidases that regulate
the coordinated elongation of glycan chains and cross-linking of stem peptides. This will allow us to
select additional drugable sites in transpeptidases and associated proteins within the PG
polymerization complexes. We will map key interactions by FRET analyses in live bacteria producing
fluorescent proteins and by in vitro transpeptidase/glycosyltransferase assays in complexes
obtained by tandem-affinity purification. Microfluidic cultures and time-lapse microscopy will assess
the impact of inhibitors on cell division and viability. The interaction of lead compounds with their
targets will be characterized by X-ray crystallography. These complementary approaches will
enable the consortium to develop novel strategies for transpeptidase inhibition and obtain leads
active against ß-lactam-resistant bacteria.
Organisations
- Newcastle University (Collaboration, Lead Research Organisation)
- UNIVERSITY OF OXFORD (Collaboration)
- University of Indianapolis (Collaboration)
- University of Amsterdam (Collaboration)
- University of Paris (Collaboration)
- Utrecht University (Collaboration)
- IBS Grenoble (Collaboration)
- UNIVERSITY OF BRITISH COLUMBIA (Collaboration)
People |
ORCID iD |
Waldemar Vollmer (Principal Investigator) |
Publications
Banzhaf M
(2020)
Outer membrane lipoprotein NlpI scaffolds peptidoglycan hydrolases within multi-enzyme complexes in Escherichia coli.
in The EMBO journal
Egan AJF
(2018)
Induced conformational changes activate the peptidoglycan synthase PBP1B.
in Molecular microbiology
Hernández-Rocamora VM
(2018)
Coupling of polymerase and carrier lipid phosphatase prevents product inhibition in peptidoglycan synthesis.
in Cell surface (Amsterdam, Netherlands)
Maya-Martinez R
(2018)
Recognition of Peptidoglycan Fragments by the Transpeptidase PBP4 From Staphylococcus aureus.
in Frontiers in microbiology
Maya-Martinez R
(2019)
Recognition of peptidoglycan fragments by the transpeptidase PBP4 from Staphylococcus aureus
in Frontiers in Microbiology
Montón Silva A
(2018)
The Fluorescent D-Amino Acid NADA as a Tool to Study the Conditional Activity of Transpeptidases in Escherichia coli.
in Frontiers in microbiology
Otten C
(2018)
Peptidoglycan in obligate intracellular bacteria.
in Molecular microbiology
Description | The Physics of Antimicrobial Resistance |
Amount | £2,158,027 (GBP) |
Funding ID | EP/T002778/1 |
Organisation | Engineering and Physical Sciences Research Council (EPSRC) |
Sector | Public |
Country | United Kingdom |
Start | 03/2019 |
End | 03/2022 |
Title | Improved technique to label bacterial cell walls |
Description | We improved the in sito labelling technique for bacterial cell wall with fluorescent D-amino acids. |
Type Of Material | Technology assay or reagent |
Year Produced | 2019 |
Provided To Others? | Yes |
Impact | The improved research tool is now available for the scientific community. |
URL | https://bio-protocol.org/e3316 |
Description | Eefjan Breukink |
Organisation | Utrecht University |
Department | Rudolf Magnus Institute |
Country | Netherlands |
Sector | Academic/University |
PI Contribution | This is a long-term scientific collaboration with Eefjan Breukink on bacterial cell wall proteins. We contribute our expertise in bacterial cell wall structure and biosynthesis, provided research tools and performed experiments. |
Collaborator Contribution | The partner contributes their expertise in peptidoglycan enzymes and precursor synthesis, provided research tools and performed experiments. |
Impact | Publications in scientific journals: PMID: 30504892 PMID: 30233559 PMID: 30093673 PMID: 30046664 PMID: 30044025 PMID: 28233869 PMID: 27709766 PMID: 27257764 PMID: 26370943 PMID: 25951518 PMID: 24821816 PMID: 22606933 PMID: 22487093 PMID: 21472954 PMID: 21183073 PMID: 17938168 PMID: 16840781 PMID: 16154998 |
Description | Jean-Pierre Simorre |
Organisation | IBS Grenoble |
Country | France |
Sector | Private |
PI Contribution | This is a long-term scientific collaboration with Jean-Pierre Simorre on bacterial cell walls and proteins. We contributed our expertise in bacterial cell wall structure and biosynthesis, provided research tools and performed experiments. |
Collaborator Contribution | The partner contributed their expertise in NMR spectroscopy for the analysis of bacterial cell walls and protein structure determination, provided research tools and performed experiments. |
Impact | Publications in scientific journals: PMID: 30713527 PMID: 30046664 PMID: 30044025 PMID: 30031884 PMID: 25951518 PMID: 24954617 PMID: 24821816 PMID: 24691651 PMID: 24493340 PMID: 22192687 PMID: 20681725 PMID: 18393418 |
Description | Jeanne Salje |
Organisation | University of Oxford |
Department | Wolfson College |
Country | United Kingdom |
Sector | Charity/Non Profit |
PI Contribution | This is a scientific collaboration with Jeanne Salje on pathogenic intracellular bacterial. We contributed our expertise in bacterial cell wall structure and biosynthesis. |
Collaborator Contribution | The partner contributed their expertise in the biology of intracellular bacteria. |
Impact | Publication in scientific journal: PMID: 29178391 |
Start Year | 2017 |
Description | Joe Gray |
Organisation | Newcastle University |
Department | Newcastle University Medical School |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | This is a long-term scientific collaboration with Joe Gray on bacterial cell walls and proteins. We contributed our expertise in bacterial cell wall structure and biosynthesis, provided research tools and performed experiments. |
Collaborator Contribution | The partner contributed their expertise in mass spectrometry for the analysis of bacterial cell walls and proteins, provided research tools and performed experiments. |
Impact | Publications in scientific journals: PMID: 31916938 PMID: 31209025 PMID: 30713527 PMID: 30102748 PMID: 30066424 PMID: 29311645 PMID: 28974693 PMID: 28581639 PMID: 27506799 PMID: 26537571 PMID: 26010014 PMID: 25424554 PMID: 24292151 PMID: 18996994 |
Start Year | 2007 |
Description | Michael VanNieuwenhze |
Organisation | University of Indianapolis |
Country | United States |
Sector | Academic/University |
PI Contribution | This is a long-term scientific collaboration with Michael VanNieuwenhze on bacterial cell wall synthesis. We contributed our expertise in bacterial cell wall structure and biosynthesis, provided research tools and performed experiments. |
Collaborator Contribution | The partner contributed their expertise in chemical probes for bacterial cell wall labelling, provided research tools and performed experiments. |
Impact | Multi-disciplinary collaboration: chemistry, microbiology Publications in scientific journals: PMID: 31916938 PMID: 31743648 PMID: 31419110 PMID: 30504892 PMID: 30275297 PMID: 30233559 PMID: 30046664 PMID: 29765094 PMID: 29311645 PMID: 28974693 PMID: 24292151 |
Start Year | 2014 |
Description | Michel Arthur |
Organisation | University of Paris |
Country | France |
Sector | Academic/University |
PI Contribution | This is a scientific collaboration with Michel Arthur on the biochemistry of cell wall enzymes based on our common interest in the bacterial cell wall. We contributed our expertise in bacterial cell wall structure and biosynthesis, provided research tools and performed experiments. |
Collaborator Contribution | The partner contributed their expertise in bacterial cell wall enzymology, provided research tools and performed experiments. |
Impact | Publication in scientific journal: PMID: 30275297 |
Start Year | 2015 |
Description | Natalie Strynadka |
Organisation | University of British Columbia |
Department | Vancouver Coastal Health Research Institute |
Country | Canada |
Sector | Academic/University |
PI Contribution | Expertise in bacterial cell wall biochemistry. Performed enzymatic assays. |
Collaborator Contribution | Expertise in structural biology. Provided protein samples. |
Impact | Scientific publication: PMID: 30713527 |
Start Year | 2015 |
Description | Tanneke den Blaauwen |
Organisation | University of Amsterdam |
Country | Netherlands |
Sector | Academic/University |
PI Contribution | This is a long-term scientific collaboration with Tanneke den Blaauwen on cell envelope biogenesis in E. coli. We contribute our expertise in bacterial cell wall structure and biosynthesis, provided research tools and performed experiments. |
Collaborator Contribution | The partner contributed their expertise in bacterial cell biogenesis and cellular localization of proteins, provided research tools and performed experiments. |
Impact | Publications in scientific journals: PMID: 30233559 PMID: 26124755 PMID: 25951518 PMID: 23387922 PMID: 22606933 PMID: 21472954 PMID: 21183073 PMID: 20545860 PMID: 16803586 |
Description | Soap box science 2018 |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Public/other audiences |
Results and Impact | Public presentation of scientific topics at the Monument city centre of Newcastle upon Tyne, called soap box science, by women. Daniela Vollmer (Research Technician in the group) participated in this event. |
Year(s) Of Engagement Activity | 2018 |
URL | http://soapboxscience.org/ |