Biochemistry & Biophysics projects

Please try to identify a research project that suits you.  This is to ensure a proper match between your research interests and your prospective supervisor. We do not encourage applications to more than one project however we would encourage informal discussions with different project supervisors to help you decide which project to apply to. Contact details for academic staff can be found on each project description.

Life pushed to extremes: probing chromosome segregation in thermophilic Archaea. Supervisors: Dr Daniela Barilla and Dr James Chong

Archaea are remarkable objects of investigation due to their exquisitely unusual biological properties and macromolecules. They are widely disseminated in the most disparate environmental niches and present unique molecular adaptations to life pushed to extremes. Thermophilic archaea are important for fundamental studies on evolution and the origin of life: they can be considered as a ‘time capsule’ providing a unique glimpse of what life was like on Earth when this was a planet bursting with geological activities billions of years ago. In addition, the molecular adaptations to life in extreme environments make archaea interesting objects of investigation for biotechnology and synthetic biology applications.

Since their discovery almost four decades ago, there has been an escalation in knowledge, genome sequences and publications on this domain of life. However, the fundamental process of chromosome segregation remains a black box in this branch of the tree of life. We have recently published the identification and initial characterization of the first chromosome segregation system in archaea [Proc
Natl Acad Sci USA 2012, 109: 3754-3759]. This genome partition machine from the archaeon Sulfolobus solfataricus consists of two proteins named SegA and SegB and a cis-acting DNA region. Our biochemical and genetic data have indicated that the SegAB complex fulfils a crucial role in chromosome segregation and is the prototype of a DNA partition machine widespread across archaea. Sulfolobus represents an excellent model system to study chromosome partition, as it harbours one single chromosome copy, in contrast to other members of the archaea domain that are highly polyploid.

Now we want to widen our investigations to dissect an orthologous system in Sulfolobus acidocaldarius. Our key hypothesis is that the products of the identified genes are involved in chromosome segregation and constitute the core components of a genome partition system. This project will test the above hypothesis and explore the molecular mechanisms underpinning chromosome segregation in archaea by interlocking, multidisciplinary approaches ranging from genetics and biochemistry to cutting-edge super resolution microscopy and single-molecule experiments. The overarching aim is to produce a detailed, mechanistic map of a fundamental biological process in a group of organisms with a remarkable and yet underexploited potential for biotechnology and synthetic biology.

Supervisors: Dr Daniela Barilla & Dr James Chong

To discuss your suitability for this project please email: daniela.barilla@york.ac.uk

Please read the 'How to apply' tab before submitting your application.

Revealing IGFN1 functional roles through in vivo CRISPR/CAS targeting and in vitro mechanical protein unfolding Supervisors: Dr Gonzalo Blanco & Dr Christoph Baumann

This project will evaluate the implication of IGFN1 in skeletal muscle mechanotransduction and selective autophagy by: 1) Generating an Igfn1 loss of function allele in single adult muscles by CRISPR/CAS mediated genome editing; 2) Measuring the elastic properties of purified IGFN1 by atomic force microscopy. Cytoskeletal protein crosslinkers have been recently placed at the forefront of cellular mechanotransduction. When mechanical strain causes these linkers to unfold, it is proposed that an inducible selective autophagy mechanism is triggered to dispose of damaged proteins. This pathway contributes to protein quality control and is essential for maintaining muscle integrity. However, the role of this pathway in processing the variety of cytoskeletal protein crosslinkers that are required to maintain the sarcomere is not clear. IGFN1 is a large muscle-specific protein conserved in all sequenced mammals with a typical domain composition similar to other proteins that organize and support the sarcomere structure by crosslinking the cytoskeleton. IGFN1 localizes to the Z-disc and is therefore a strong candidate for a mechanotransduction sensor in skeletal muscle. The Igfn1 gene will be targeted using CRISPR/CAS customized vectors previously validated in vitro to generate loss of function mutations in hindlimb muscles of adult mice in vivo. The loss of function effects on muscle structure and function will be characterized in detail by histology, immunofluorescence with autophagy markers and electron microscopy. The lengths obtained by AFM will be related to the physical dimensions of the sarcomeric structures during the cycles of contraction and relaxation. These experiments will provide mechanistic insights into the role of IGFN1 as a structural stabilizer and client of selective autophagy.

Supervisors: Dr Gonzalo Blanco & Dr Christoph Baumann

To discuss your suitability for this project please email: gonzalo.blanco@york.ac.uk

Please read the 'How to apply' tab before submitting your application. 

Single-molecule investigations of antibacterial proteins. Supervisors: Dr Christoph Baumann & Prof Marek Brzozowski

This project will focus on the nuclease colicins, which are protein antibiotics that parasitize essential outer membrane (OM) and periplasmic proteins to gain entry into Escherichia coli cells. The import of a single enzymatic colicin molecule into a bacterial cell is sufficient to induce cell death. Therefore, a single-molecule approach to investigating the mechanism used by a colicin molecule to cross the cell envelope is biologically relevant. While many of the molecular components for cell entry have been characterised, it remains unknown how these systems expedite translocation into cells. This project will provide fundamental knowledge about the mechanism whereby species-specific antibacterial proteins translocate into Gram-negative bacteria, and the molecular organisation of the bacterial cell envelope. The student will learn and apply novel biochemical and single-molecule fluorescence techniques to follow colicin cell binding and import. The microscopy techniques include single-molecule tracking, total internal reflection fluorescence (TIRF) microscopy, fluorescence recovery after photobleaching (FRAP), and single-molecule fluorescence resonance energy transfer (smFRET). These investigations will be guided by recent structural insight into the OM translocation mechanism used by nuclease colicins. The knowledge gained from this project will greatly improve our understanding of bacterial physiology. This knowledge will be invaluable in the search for new species-specific antibiotics at a time when the number of antibiotic-resistant microbes is increasing. This project is part of an ongoing collaboration with Prof. Colin Kleanthous (University of Oxford), an expert in the molecular enzymology of colicins.

Supervisors: Dr Christoph Baumann (Biology) & Prof Marek Brzozowski (YSBL)

To discuss your suitability for this project please email: christoph.baumann@york.ac.uk

Please read the 'How to apply' tab before submitting your application.

Microbially Catalysed Electrolytic H2 Production. Supervisors: Dr James Chong & Dr Alison Parkin

Harnessing microbes cultured on waste water as fuel-producing catalysts is a tantalizing prospect: unlike isolated-enzyme systems, living cells retain the ability to regenerate their active site centres, providing the potential to construct devices with practically-useful, long lifetimes from minimal resources. We have recently demonstrated the ability of methanogenic microbes to catalyse electrolytic H2 production with minimal energy input.

This highly interdisciplinary project aims to understand and optimize methanogen-based H2-production by exploring the potential that the unique range of seven hydrogenases found in these organisms offers for efficient electricity storage. Using a complementary suite of techniques working across the chemical-biochemical-biological interface, experiments will focus on:
· increasing the rate and density of gas production through synthesis of complementary biological-electrode surface chemistry
· integrating hydrogen extraction with downstream biobased chemical production
· screening of microbial communities for selection of an optimized methanogen chassis

The student undertaking this project will be exposed to electrochemical measurements, gas chromatography, anaerobic microbiology and molecular biology for the genetic manipulation of methanogens.

Supervisors: Dr James Chong (Biology) & Dr Alison Parkin (Chemistry)

To discuss your suitability for this project please email: james.chong@york.ac.uk

Please read the 'How to apply' tab before submitting your application.

 

 

Nuclear matrix support of DNA replication and transcription Supervisors: Dr Dawn Coverley & Prof Bob White

Visualization of the structure of nuclei has been the main method of cancer diagnosis since light microscopy allowed. However, very little is known about the underlying molecular alterations that give rise to aberrant nuclear architecture, or the effect these have on the organisation of nuclear processes like DNA replication and transcription. Some proteins that are involved in the regulation of DNA replication are immobilized by attachment to the nuclear matrix in normal cells, but not in cancer cells (or in undifferentiated cells). This project will ask whether the same is true for proteins that regulate transcription, in order to test the extent to which published observations (1) can be generalised beyond DNA replication. The project will also look in detail at the CIZ1 protein to identify the proteins that it normally interacts with in the nuclear matrix. Unlike the cytoskeleton, there is currently no consensus on the main protein components of the nuclear matrix, making this approach particularly interesting.

CIZ1 has been implicated in a several common human cancers as well as chronic age-related disorders, and is being developed as the basis for a blood test for cancer (2). Its interaction partners may be similarly useful as markers of nuclear integrity (for application in regenerative medicine strategies or analysis of cancer cells), and might offer new therapeutic drug targets. There are very few molecules that can be used for function-related studies of the nuclear matrix making this work both novel and timely. The project will involve mammalian cell culture and cell cycle synchrony techniques, sub-cellular fractionation to isolate nuclear matrix fractions using a range of techniques, nuclear stability assays to compare the integrity of the nucleus in different cell types, immunofluorescence microscopy and related image quantification techniques, and protein interaction studies including 2D-LC-MS to discover new components of the nuclear matrix.

Supervisors: Dr Dawn Coverley & Prof Bob White

To discuss your suitability for this project please email: dawn.coverley@york.ac.uk

Please read the 'How to apply' tab before submitting your application.

Structure and Function of HTLV-1 Transcriptional Regulators. Supervisors: Dr Fabiola Martin, Prof Robert White & Dr Fred Antson

HTLV-1 is a virus that incorporates itself into human DNA and can be transmitted through unprotected sex, breastfeeding and contaminated blood transfusions.  An estimated 15-20 million people are infected worldwide, of whom 2-5% develop the aggressive blood malignancy ATLL, which has an average survival time of only nine months.  Another 3% of HTLV-1 carriers develop severe walking disability, which progresses painfully and irreversibly to wheelchair-dependence.  HTLV -1 infection is a neglected disease, for which there is no vaccine or cure.

ATLL is thought to be caused by two proteins produced by the virus in human cells, Tax and HBZ.  We aim to analyse these viral proteins to gain insight into how they function.  This will involve characterising some of the changes they induce in expression of host non-coding RNAs and the mechanism(s) of such changes.  We will also investigate the structures of these key viral proteins, which could be very helpful in understanding how they operate.  Our studies aim to provide detailed insight, at the molecular level, into the properties of the viral proteins that cause malignancy, an important step towards tackling this scourge.   Thorough training will be provided in in biochemistry, structural and molecular biology, geared towards systems of clinical relevance.

Supervisors: Dr Fabiola Martin (Biology/HYMS), Prof Robert White (Biology) & Prof Fred Antson (YSBL)

To discuss your suitability for this project please email: fabiola.martin@hyms.ac.uk

Please read the 'How to apply' tab before submitting your application.

Characterization of the LIMD1-VHL-PHD2 complex and its role in the hypoxic response Supervisor: Dr Michael Plevin, Dr Dimitris Lagos & Dr Tyson Sharp

Multi-cellular organisms have tightly regulated systems of oxygen homeostasis to ensure that cells and tissues are supplied with sufficient oxygen. In humans, each tissue type requires a certain concentration of oxygen, below which essential cellular processes such as energy production, protein synthesis and cell growth and division become impaired. Rapid reaction and adaptation to low oxygen concentrations ('hypoxia') can reduce damage and enable cells to remain viable. This process is known as the 'hypoxic response'. Dysfunction of the hypoxic response is implicated in many human diseases, including chronic anaemia and ischemia, as well as in the growth and spread of cancers.

The molecular basis for hypoxic signalling has been mapped out: in low oxygen conditions, a family of hypoxia-inducible transcriptions factors (HIFs) control the expression of various factors that facilitate the hypoxic response. However, under normoxic conditions, specific proline residues in one of the HIF subunits (HIF-α) are hydroxylated by one of 4 prolyl hydroxylases (PHDs). These hydroxyproline modifications are recognised by von Hippel Landau protein (VHL), which recruits an E3 ligase complex, leading to ubiquitination of HIF and its eventual degradation.

A 676 amino acid scaffold protein called LIMD1 has recently been shown to be critical to the hypoxic response signalling pathway. This discovery represents a new level of molecular biology in hypoxia signalling. The objective of this project is to determine how the structure-function relationship of LIMD1 mediates its affect on hypoxic signalling. We will employ a range of structural and biophysical approaches to characterise the structure and dynamics of LIMD1 and explore how it interacts with other components of the HIF signalling pathway, including VHL and PHDs. These molecular level analyses will be complemented by cell‐based assays of hypoxic signalling.

Deregulation of the hypoxic response is a key characteristic in cancer development, large tumour growth and the spread of cancer throughout the body. Furthermore, dysfunction of this key cellular control process is linked to many non‐cancerous diseases, such as neurological disease, myocardial infarction (heart attacks), stokes, and many ischemic (low oxygen) related diseases. The molecular-level analyses of LIMD1 proposed here will drive studies into small molecule modulation of hypoxic signalling.

Supervisor: Dr Michael Plevin (Biology), Dr Dimitris Lagos (Biology) & Dr Tyson Sharp (Centre for Molecular Oncology, Queen Mary University of London)

To discuss your suitability for this project please email: michael.plevin@york.ac.uk

Please read the 'How to apply' tab before submitting your application.

 

How a bacterium spins a web: structural and functional studies on the biosynthesis of an adhesive surface polymer in uropathogenic Escherichia coli Supervisors: Prof Jennifer Potts & Dr Gavin Thomas

Multidrug-resistant Escherichia coli, which cause life-threatening bloodstream infection (bacteremia), are emerging as a major international healthcare risk. These bacteria almost always reach the bloodstream as a result of urinary tract infection (UTI). Prevention of E. coli colonization of the urinary tract (or urinary catheters) could reduce systemic infection and this is emerging as an important route for intervention. A recent study using an E. coli UTI model highlighted a number of genes required for colonization including genes involved in the biosynthesis of polymeric N-acetylglucosamine (PNAG). PNAG is a cell surface located polymer, already known to be a major factor in the colonization of catheters. The aim of the project is to study the structure and function of PgaA, an outer-membrane protein that acts as a transporter for PNAG. PgaA contains both periplasmic (soluble) and membrane embedded domains; both will be studied. This is a collaboration between a structural biology (Potts) and Microbiology (Thomas) lab which occupy adjacent lab space. A wide range of techniques, for example X-ray crystallography, NMR spectroscopy, surface plasmon resonance and isothermal titration calorimtery) are available to study the structure and ligand binding of PgaA. Hypotheses regarding key regions of the protein required for functional interactions will be tested through in vivo mutation in E. coli.

Supervisors: Prof Jennifer Potts & Dr Gavin Thomas 

To discuss your suitability for this project please email: jennifer.potts@york.ac.uk

Please read the 'How to apply' tab before submitting your application. 

Eating the poison: characterization of the molecular basis of antimicrobial peptide resistance in pathogenic Escherichia coli. Supervisors: Dr Gavin Thomas & Prof Tony Wilkinson

Cationic antimicrobial peptides (CAMPs) are an important first line of defence in animals, which kill bacteria by forming pores in their membranes. One important mechanism by which Gram –ve pathogens have evolved resistance to CAMPs is to prevent their incorporation into membranes by recognizing and capturing them in the periplasm and transporting them into the cell for degradation. This is the function of the Sap ABC transporter system which was first discovered in Salmonella but is important in other pathogens like Escherichia coli and Haemophilus infleunzae. While SapA, the periplasmic substrate binding protein (SBP) of the Sap system must bind CAMPs to enable their uptake, the substrates for this SBP are atypical of ABC peptide transporters being over 10-fold larger than the normal di- or tri-peptides that these transporters recognise. There are no structural data for SapA proteins and consequently how they recognize the CAMPs, which are often over 35 residues in length, must involve a significantly different ligand binding mechanism to other peptide binding SBPs. Building on the in-depth expertise in ABC transporter structure & function in York, our main objectives are to take a primarily biochemical and structural approach to understand how SapA has evolved from its close relative DppA (a dipeptide SBP) to recognise and transport a wide range of long and potentially folded peptides. Both supervisors have long track records of working together with SBPs both in vitro and in vivo (see key publications) and the student will be trained in a wide range of structural (X-ray crystallography) and biochemical methods in this jointly supervised project.

Supervisors: Dr Gavin Thomas (Biology) & Prof Tony Wilkinson (Chemistry)

To discuss your suitability for this project please email: gavin.thomas@york.ac.uk

Please read the 'How to apply' tab before submitting your application. 

The biological role and structure function analyses of O-antigen modifying enzymes in the bacterial pathogen Salmonella Supervisors: Dr Marjan van der Woude & Dr Gavin Thomas

Salmonella is an important bacterial pathogen, affecting both humans and livestock. One virulence strategy Salmonella uses is to modify its cell surface structures to facilitate immune evasion or directly enhance virulence. This project focuses on how and why Salmonella modifies the O-antigen part of its lipopolysaccharide (LPS). The O-antigen plays a pivotal role as a virulence factor, immunodominant antigen, and phage receptor, and may influence pathogen-host tissue interactions. Modifications are thus likely to impact many aspects of this bacterial pathogen’s biology. This project builds upon a body of work in from the supervisor’s lab and concerns one specific family of O-antigen modifying proteins with a predicted acetyltransferase function. Using a broad range of molecular biology, genetic and biochemistry approaches you will study the structure/function of these proteins and you will examine the role of the modification in a set of biological assays.

The project will further our understanding of this family of little studied proteins and the process of O-antigen modification, contribute to our understanding of Salmonella as a pathogen and inform the development of LPS based vaccine approaches.

Techniques you will use include but are not limited to mutagenesis, cloning, protein expression and purification, electrophoresis, and use of software for sequence analysis/protein structure. Lab meetings, seminar series, minisymposia and the opportunity to participate in and develop outreach activities will help you develop a broad range of scientific and transferable skills. There is a vibrant PhD community that hosts events like Coffee and careers. You will be co-supervised and will carry the work out in labs specializing in protein biochemistry as well as molecular microbiology, housed in the Centre of Immunology and Infection, Department of Biology and YSBL. This project is suitable for applicants with a strong background and interest in microbiology, biochemistry and/or molecular biology, and an overall interest in the molecular basis of bacterial pathogenesis.

Supervisors: Dr Marjan van der Woude & Dr Gavin Thomas

To discuss your suitability for this project please email: marjan.vanderwoude@york.ac.uk

Please read the 'How to apply' tab before submitting your application.