Interaction and interdependency of GlmM and DacA - two essential enzymes required for methicillin resistance in Staphylococcus aureus
Lead Research Organisation:
Imperial College London
Department Name: Infectious Disease
Abstract
Staphylococcus aureus is frequently found on the skin, nares or other mucosal surfaces of healthy individuals without causing any symptoms or harm. However, S. aureus can also cause a wide variety of infections ranging from small skin infections to life-threatening invasive diseases. Alarmingly, many S. aureus strains have acquired resistance to multiple antimicrobials and as a result S. aureus is still one of the leading causes of antibiotic-resistant healthcare-associated infections worldwide. The resulting difficulty in treating these infections is a major public health concern. Therefore, it is of interest to study biosynthesis pathways and enzymes that are required for bacterial growth to ultimately develop new antimicrobials. The purpose of this study is to characterize the interaction and interdependency of the two staphylococcal enzymes GlmM and DacA. Both enzymes are extremely important for the growth of S. aureus. At the first glance there is no apparent connection between them as GlmM is required for the production of an essential cell wall precursor and DacA is responsible to the production of a recently discovered signalling nucleotide c-di-AMP. However, recent work has indicated that there is a direct protein/protein contact between these two enzymes, suggesting that the pathways that they regulate are somehow interconnected. DacA and GlmM are therefore potentially central players in coordinating the synthesis of the cell wall polymer peptidoglycan with pathways that are regulated by c-di-AMP. The aims of this project are to provide an image of the DacA/GlmM complex in order to show exactly how these two proteins interact, find out how the activity of one enzyme influences the activity of the other and how this interaction affects the growth and methicillin resistance of the cell. In summary, the work will yield valuable information on the interdependency of two important bacterial enzymes, which interconnect two essential cellular processes in the Gram-positive bacterial pathogen S. aureus. Due to the direct link of S. aureus to human infection, the output of this work on essential cellular pathways can potentially offer future opportunities for the development of novel therapeutic approaches interfering with the growth of S. aureus and potentially a number of other Gram-positive pathogens.
Technical Summary
The cell wall in Staphylococcus aureus is composed of a thick peptidoglycan layer, cell surface proteins, teichoic acids and carbohydrate polymers. Proper assembly of the different structures is not only essential for the pathogenesis of S. aureus but also for bacterial growth and several antimicrobial agents target the synthesis of key cell wall components. GlmM is a phosphoglucosamine mutase required for the production of UDP-N-acetylglucosamine, an essential cytoplasmic precursor for the synthesis of peptidoglycan. In S. aureus and many other Gram-positive bacteria, glmM forms a three-gene operon with dacA and ybbR, which encode the c-di-AMP synthase DacA and the membrane protein YbbR that is thought to regulate the activity of cylcase, respectively. c-di-AMP is a recently discovered signalling molecule that is essential for growth of S. aureus and many other Gram-positive bacteria in standard laboratory conditions. Recent work has highlighted a physical interaction between GlmM and DacA, suggesting that these enzymes are central players in coordinating peptidoglycan synthesis with the metabolic pathways that are regulated by c-di-AMP. These include in S. aureus potassium and osmolyte uptake as well as pH homeostasis. The aims of this project are to characterize the interdependency of the DacA and GlmM enzyme activities using in vitro assay systems, to determine the structure of the DacA/GlmM complex, to investigate the in vivo dynamics and the functional consequence of the DacA/GlmM interaction on the metabolism and methicillin resistance of S. aureus and to conclusively determine if GlmM is essential for the growth of S. aureus to resolve conflicting data. All together this work will allow us to propose a model for the molecular mechanism by which binding of GlmM can influence the activity of the c-di-AMP cyclase, provide insight into the biological significance of this protein interaction and uncover new avenues to interfere with the growth of S. aureus.
Planned Impact
For details please see pathways to impact document. In brief, the outcomes of this research project will provide information on the structures and molecular basis by which two central Staphylococcus aureus enzymes, DacA and GlmM, interact to coordinate the essential cellular processes of c-di-AMP signalling and peptidoglycan synthesis. Both enzymes are conserved and present in a large number of other Gram-positive bacteria and hence information gained from this work will have an impact on a variety of different academic research fields. Many S. aureus strains have acquired resistance to multiple antimicrobials and S. aureus remains a leading cause of antibiotic-resistant healthcare-associated infections worldwide. The resulting difficulty in treating these infections is a major public health concern. Therefore, it is of interest to study biosynthesis pathways and enzymes such as described in this application that are required for bacterial growth to ultimately develop new antimicrobials. Therefore, this work will also be of interest to pharma as well as health care professionals.
Organisations
- Imperial College London (Lead Research Organisation)
- National Center for Scientific Research (Centre National de la Recherche Scientifique CNRS) (Collaboration)
- Eberhard Karls University of Tübingen (Collaboration)
- University College London (Collaboration)
- ETH Zurich (Collaboration)
- IMPERIAL COLLEGE LONDON (Collaboration)
Publications
Chee Wezen X
(2022)
Structure-Based Discovery of Lipoteichoic Acid Synthase Inhibitors.
in Journal of chemical information and modeling
Chee Wezen X
(2022)
Structure-Based Discovery of Lipoteichoic Acid Synthase Inhibitors.
Chee Wezen X
(2022)
Structure-Based Discovery of Lipoteichoic Acid Synthase Inhibitors.
Pathania M
(2021)
Structural basis for the inhibition of the Bacillus subtilis c-di-AMP cyclase CdaA by the phosphoglucomutase GlmM
in Journal of Biological Chemistry
Pompeo F
(2018)
Investigation of the phosphorylation of Bacillus subtilis LTA synthases by the serine/threonine kinase PrkC.
in Scientific reports
Rismondo J
(2021)
EslB Is Required for Cell Wall Biosynthesis and Modification in Listeria monocytogenes.
in Journal of bacteriology
Rismondo J
(2020)
GtcA is required for LTA glycosylation in Listeria monocytogenes serovar 1/2a and Bacillus subtilis.
in Cell surface (Amsterdam, Netherlands)
Rismondo J
(2021)
Modifications of cell wall polymers in Gram-positive bacteria by multi-component transmembrane glycosylation systems.
in Current opinion in microbiology
Title | Bacterial strains |
Description | A number of bacterial stains were created as part of this research and have now been published. The strains will be shared with the research community upon request |
Type Of Material | Biological samples |
Year Produced | 2017 |
Provided To Others? | Yes |
Impact | Not yet applicable |
Description | Anne Galinier |
Organisation | National Center for Scientific Research (Centre National de la Recherche Scientifique CNRS) |
Country | France |
Sector | Academic/University |
PI Contribution | We provided expertise and performed experiments to analysis Lipoteichoic acid production in wild-type and mutant Bacillus subtitles strains |
Collaborator Contribution | The collaborator was the lead group in this project |
Impact | Publication |
Start Year | 2017 |
Description | Anne Galinier |
Organisation | National Center for Scientific Research (Centre National de la Recherche Scientifique CNRS) |
Country | France |
Sector | Academic/University |
PI Contribution | We provided expertise and performed experiments to analysis Lipoteichoic acid production in wild-type and mutant Bacillus subtitles strains |
Collaborator Contribution | The collaborator was the lead group in this project |
Impact | Publication |
Start Year | 2017 |
Description | Christoph Mayer |
Organisation | Eberhard Karls University of Tübingen |
Country | Germany |
Sector | Academic/University |
PI Contribution | We were a collaborator on this project and purified and provided bacterial lipoteichoic acid cell wall material for experiments performed by the lead research team |
Collaborator Contribution | The collaboration parter was the lead authors on this work |
Impact | Publication |
Start Year | 2019 |
Description | Marc Dionne |
Organisation | Imperial College London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | Our group was the lead on this work - conception, experimentation, data analysis and manuscript writing |
Collaborator Contribution | Expertise and work performed for a Drosophila/Listeria infection experiment |
Impact | Publication |
Start Year | 2019 |
Description | Martin LOESSNER |
Organisation | ETH Zurich |
Country | Switzerland |
Sector | Academic/University |
PI Contribution | We performed an NMR analysis of LTA isolated from WT and mutant Listeria monocytogenes strains We hosted a student for a few month from the collaboration lab |
Collaborator Contribution | Mass spec analysis of WTA |
Impact | 1 Publication and two further publications have been submitted |
Start Year | 2018 |
Description | Native mass spec |
Organisation | University College London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | We provided a protein samples for a native mass sept experiment to determine size of a protein complex. |
Collaborator Contribution | A Phd student in the partners lab analysed the size of the protein complex by native mass spectrometry. The know how and equipment to perform such an experiment was provided. The obtained data formed part of a recent collaborative publication |
Impact | PLOS Pathogen publication Accepted in January 2019 |
Start Year | 2018 |
Description | CMBI Superbug Zone at the Imperial Festival |
Form Of Engagement Activity | Participation in an open day or visit at my research institution |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Public/other audiences |
Results and Impact | Stalls presenting ongoing research, mock labs, microscopy stall |
Year(s) Of Engagement Activity | 2018 |
Description | US Undergraduate student visit - October 2019 |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Undergraduate students |
Results and Impact | Arranged visit for ca 15 undergraduate student from the US currently undertaking a semester abroad to our Centre. As part of the visit, the students got lab and facility tours, an overview about the activities going on in our Centre, and short presentations by Students and post docs about "life as a PhD student and post doc" and what to expect |
Year(s) Of Engagement Activity | 2019 |
Description | University of Leicester - Undergraduate student visit - April 2019 |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Undergraduate students |
Results and Impact | Arranged visit for ca 25 undergraduate student from the University of Leicester to our Centre. As part of the visit, the students got lab and facility tours, an overview about the activities going on in our Centre, and short presentations by Students and post docs about "life as a PhD student and post doc" and what to expect |
Year(s) Of Engagement Activity | 2019 |