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Understanding the role of protein mechanical strength in biofilm formation and function

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  • Full or part time
    Dr D Brockwell
  • Application Deadline
    No more applications being accepted
  • Funded PhD Project (European/UK Students Only)
    Funded PhD Project (European/UK Students Only)

Project Description

A PhD studentship (based in Leeds but co-supervised by Dr William Durham, Department of Physics and Astronomy, University of Sheffield) is available as part of a White Rose Studentship Network funded directly by Universities of Leeds, Sheffield and York. The network has 3-linked studentships (the others in York and Sheffield) with the overall theme of “Bacterial surface proteins: linking biophysics to biology”. The network will examine molecules (protein domains) to systems (biofilms) approaches through an inter-institutional collaboration of experts in structural biology, biophysics and microbial science.

The Leeds-based project: Understanding the role of protein mechanical strength in biofilm formation and function

Many proteins resist or respond to mechanical deformation as part of their function. The importance of understanding how mechanical force affects both the structure and the binding of proteins to each other and ligands / surfaces is exemplified by biofilm formation, which often occurs under hydrodynamic stress (i.e. in the vasculature). We have shown previously that a repetitive surface protein involved in host adhesion and bacterial-bacterial interactions (SasG) essential for biofilm formation displays extreme mechanical stability (Gruszka et al. Nature Commun (2015)). However, many questions remain unanswered that, if addressed, may lead to novel methods of preventing or removing biofilms. These include: why are these proteins so strong, how do they dimerise to form bacterial interactions, what happens if the strength of this interaction under force is weakened, what is the effect of destabilising these proteins (or their dimerization) on biofilm formation ‘in vivo’?

Objectives
1. The effect on the mechanical strength of SasG of introducing β-strand-, hydrophobic- or charge-disrupting mutations will be assessed using force spectroscopy.
2. Variants with interesting force characteristics will be introduced into S. aureus to reveal their effect on biofilm formation. Custom microfluidic devices and three-dimensional confocal imaging will be used to quantify surface attached biomass and biofilm morphology. Carried out in Sheffield
3. Dimerization of SasG is necessary for biofilm formation but the mechanism, the role of metals and the effect of force are still unclear. We will perform ‘fly-fishing’ experiments to measure the formation and rupture force of the complex and the effects of metal ions and / or specific amino-acid substitutions.
4. We will select conditions / variants with modulated dimer properties and assess their effects in vivo. We will leverage microfluidic systems and automated image analyses, to understand how dimer destabilization affects the initialization of biofilms under physiologically relevant hydrodynamic conditions. Carried out in Sheffield

The student will be in well-funded laboratories with all equipment needed. The School of Molecular and Cellular Biology / Faculty of Biological Science at Leeds has world-class facilities that include two Atomic Force Microscopes that are optimised for force spectroscopy experiments (Asylum Research, MFP-3D).
Microfluidic experiments will be conducted at the University of Sheffield, where the necessary cell culturing, microfluidic device fabrication, and automated time lapse imaging facilities are available.

The proposed project is highly multi-disciplinary, combining biophysics, biochemistry and microbiology. The student will gain training in single molecule force spectroscopy, protein/lipid chemistry, surface functionalisation, microfluidic assays, high throughput image analysis, and computational techniques together with microbiological methods.

Funding Notes

4 year White Rose Research Studentship 2017/18
The successful applicant will receive fees and stipend (c.£14,296). The PhD will start in Oct 2017. Applicants should have, or be expecting to receive, a 2.1 Hons degree in a relevant subject.
There are two stages to the application process:
1) apply for a PhD position via the University of Leeds online application https://studentservices.leeds.ac.uk/pls/banprod/bwskalog_uol.P_DispLoginNon
2) apply for the White Rose Network scholarship via http://www.leeds.ac.uk/rsa/postgraduate_scholarships/wrs-2350

References

Cooperative folding of intrinsically disordered domains drives assembly of a strong elongated protein. Gruszka,. D., Whelan, F., Farrance, O., Fung, H., Paci, E., Jeffries, C., Svergun, D., Baldock, C., Baumann, C., Brockwell, D., Potts, J. and Clarke, J. (2015) Nat Commun 6: article no. 7271.

How good is research at University of Leeds in Biological Sciences?

FTE Category A staff submitted: 60.90

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