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DPhil Studentship Available

4-YEAR EPSRC CASE Studentship: Multiscale Molecular Simulations to Probe Lipid Bilayer Modifications Causing Antimicrobial Resistance

Department of Biochemistry, University of Oxford

Industrial Sponsor: IBM

Supervisors: Professor Mark Sansom, Dr Maike Bublitz & Dr Phill Stansfeld

See http://www.bioch.ox.ac.uk/graduate/postgraduateresearchstudentships

Please quote Studentship Source Code: EPSRC1

PROJECT DESCRIPTION

Molecular simulations allow us to probe the physical properties of increasingly complex and bio-realistic models of cell membranes. This will allow us to probe the role of lipid modifications to bacterial membranes which render bacteria resistance to ‘last defence’ antibiotics such as daptomycin and to new generation antimicrobial peptides (AMPs). Multiscale molecular dynamics simulations will be used to explore lipids bilayers of various compositions, analysing their physical properties and calculating free energy landscapes for interaction of antibiotics and related compounds with wild type vs. modified (i.e drug resistant) membranes. This studentship will involve a collaboration between the groups of Mark Sansom and Phillip Stansfeld (Oxford Biochemistry), who have expertise in multiscale modelling of complex cell membranes, of Maike Bublitz (Oxford Biochemistry), who conducts experimental biochemistry and structural biology of cell membranes and their proteins, and IBM Research, Hartree Centre, with expertise in Big Data and cognitive systems approaches to molecular simulations. The project will provide the student with training in molecular simulations and advanced approaches to analysis of large simulation datasets. There will also be opportunities for exposure to and collaboration with experimentalists in bacterial membrane systems. The outcome will be simulation toolkit for analysis of antibiotic interactions with complex models of bacterial membranes, providing physical insights into mechanisms of current and future antimicrobial resistance.

Students with a chemistry, biophysics or biochemistry background, ideally with some relevant undergraduate computational experience, would be preferred.

Informal enquiries are welcome: mark.sansom@bioch.ox.ac.uk, phillip.stansfeld@bioch.ox.ac.uk & maike.bublitz@bioch.ox.ac.uk.

The project is supported by a 4-year CASE PhD studentship covering fees at home/EU rate plus a living allowance of not less than £14,777 per annum plus an additional supplement paid by the industrial sponsor.

To apply for this EPSRC-funded studentship, please submit an online application to the University of Oxford for admission to the D.Phil. in Biochemistry (course code: RD_BC1) by the deadline 12.00 noon (UK time) 13th April 2018. It is very important that you quote Studentship Source Code EPSRC1. No research proposal is required as part of the application. Instead you are required to upload a personal statement no more than 1000 words, describing your motivation and aptitude for this position, and your CV. Please arrange that three referees directly submit references for you.

Eligibility: ONLY students who have established UK residency are eligible to apply. For further details about residence requirements follow link:

https://www.epsrc.ac.uk/skills/students/help/eligibility/

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Visualising Targets for Novel Antibiotics

Scientists have taken significant strides towards the development of new antibiotics in our battle against resistant strains of bacteria. Researchers at Oxford, Trinity College Dublin, Diamond and the University of East Anglia have visualised, in high definition, proteins that are crucial to bacterial survival.

Indeed one of these proteins is so important that bacteria designed a natural antibiotic, globomycin, as a weapon to kill its competitors, by binding to this protein.

LspA-cover
Globomycin inhibits LspA

The protein in question is LspA: a key enzyme in both gram-negative bacteria, like E. coli, where resistance is rapidly on the rise, and gram-positive bacteria like the infamous MRSA and and multi-drug resistant tuberculosis.

Martin Caffrey’s group at Trinity College Dublin, in collaboration with Dr Phill Stansfeld in the Department of Biochemistry, University of Oxford, have perceived and studied how globomycin binds to LspA.

The study, which is published in the journal Science, provides a means to design new drugs that are similar to globomycin, thereby paving the way for novel antibiotics to kill the resistant and infectious bacteria in the future.

Lipoprotein-pathway
The Lipoprotein Pathway

Globomycin works by preventing the correct development of ‘bacterial lipoproteins’. These proteins perform critical roles in behaviour, pathogenicity, and antibiotic resistance of bacteria. Processing of these proteins requires the LspA enzyme to cut off part of the protein, whilst other enzymes stick the proteins to the outside of the cell through fatty anchors.

In bacteria, such as E. coli, the lipoprotein complex called LptDE is the bacteria’s bricklayer and is essential for the formation of the fortifications of the bacterial cell. The first structure of this complex was reported in Nature in 2014 by Chanjiang Dong’s group, at University of East Anglia, with Dr Phill Stansfeld a key component of the collaboration.

BamABCDE
The BAM Complex

The same team have now visualised and assessed another essential lipoprotein complex, called BAM, which is also reported in Nature. This protein is crucial to the building of proteins that permit the entry of nutrients into the bacterial cell, and also pump out hazardous compounds, such as antibiotics.

Disruption of this complex provides another route to the development of new antimicrobial agents.

LspA – the processing enzyme for lipoproteins, inhibited by the antibiotic Globomycin:
http://science.sciencemag.org/content/351/6275/876

BAM – the protein responsible for inserting proteins into the bacterial outer membrane:
http://www.nature.com/nature/journal/vaop/ncurrent/abs/nature17199.html

LptDE – the bacterial bricklayer:
http://www.nature.com/nature/journal/v511/n7507/full/nature13464.html

Structure & Simulations of Antibiotic Inhibition of a Signal Peptidase

Lipoproteins make up only a small fraction of a bacteria’s genome, however they perform critical roles in bacterial physiology, pathogenicity, and antibiotic resistance. Their roles include modulation of the cell envelope structure, signal transduction and transport. Like the majority of membrane proteins, Lipoproteins are targeted to the cell envelope by a  signal peptide, which affixes them to the inner bacterial cell membrane. This orients the protein so that its lipoprotein domain is in the periplasm or outside the cell. Post-translational processing of these proteins requires requires a signal peptidase II (also known as LspA) to remove the signal peptide, whilst retaining the lipoprotein’s lipid anchor.

The crystal structure of LspA from the gram-negative bacteria Pseudomonas aeruginosa has been solved by Martin Caffrey’s group at Trinity College Dublin (TCD). This 2.8 angstrom resolution structure is complexed with the antimicrobial globomycin. We have taken this structure and computationally reincorporated it back into its host membrane. In doing so we have developed molecular parameters for globomycin and simulated the inhibition of the enzyme by this natural antibiotic. We have also used molecular modelling techniques to predict how the native lipoprotein substrates of LspA engage with the enzyme, and to propose mechanisms by which the signal peptide is cleaved.

Bioinformatic analyses illustrate the conservation of a pair of aspartic acid residues, stabilised by a network of asparagine sidechains. Their importance is confirmed by mutagenesis studies, which firmly suggest LspA as an aspartyl peptidase.  In an example of molecular mimicry, globomycin appears to inhibit by acting as a noncleavable peptide that sterically blocks the active site. This structure should inform rational  drug discovery of novel antibiotics in our combat against drug resistant infections.

The link to the article is here.

Molecular Simulations of Diacylglycerol Kinase

A fantastic new structure of the integral membrane protein Diacylglycerol Kinase (DgkA) has been solved by Martin Caffrey’s group at Trinity College Dublin. This is a direct follow-up to their original study that appeared in Nature, two years previous [1]. The new structure not only shows the three-dimensional structure of the protein, but also reveals the mechanism of interaction of the substrates ATP and the lipid Monoacylglycerol within the catalytic site.

We have complemented this study by illustrating how the protein sits within a biological membrane, its molecular stability and how it dynamically interacts with substrates, products and potential molecular intermediates. More details can be found at Nature Communications [2]. To explore the structure in more detail go to the links at the PDB or MemProtMD.

References:

[1] Li, D. et al. (2013) Crystal structure of the integral membrane diacylglycerol kinase. Nature

[2] Li, D, Stansfeld, P.J. et al. (2015) Ternary structure reveals mechanism of a membrane diacylglycerol kinase. Nature Comms. 6: 10140