... understanding life in molecular detail

Dr David Brockwell

Protein, folding/unfolding, force, AFM


The Brockwell group uses single molecule force techniques and other biophysical methods to elucidate how Nature exploits mechanical force, protein structure, cooperativity and dynamics to carry out cellular function.

Current major projects include:
  • The effect of force on bacterial mechano-transduction complexes
  • The mechanical and dynamic properties of extremophilic proteins
  • Folding mechanisms of bacterial outer membrane proteins
  • Aggregation mechanisms of biopharmaceuticals

Research in the Brockwell group is carried out in well equipped modern lab space shared with the Radford creating a lively atmosphere with swapping of ideas and suggestions both through shared group meetings and general lab banter. Our Asylum Research AFM instruments are housed in the School of Physics and Astronomy at Leeds.  This allows regular contact with colleagues Simon Connell, Lorna Dougan and Neil Thomson.

Currently research in the group is following three themes which are described below. If you are interested in joining the lab to study for a PhD find studentship and eligibility details here.

 

Mechanical unfolding of proteins.

In vivo many proteins are required to resist or respond to mechanical stimuli. Over the last decade the development of atomic force microscope (AFM) instruments with high force sensitivity and sub-nanometre distance resolution has allowed the mechanical properties of single protein molecules to be measured. This work has revealed that proteins with similar stability to chemical denaturants can behave very differently when unfolded by the AFM. We are interested in finding out why some proteins are able to resist greater unfolding forces than others to understand how Nature uses mechanical denaturation to accelerate protein unfolding. In addition, we also use this information to rationally design proteins with novel mechanical properties.

Publication: Identification of a mechanical rheostat in the hydrophobic core of protein L.  Sadler et al. (2009) J Mol Biol 393:237-248.   

Properties of extremophilic proteins.

Life is found in the harshest of environments: low and high temperatures (< 0 and > 100 °C, respectively), low pH (< pH 2) and high salt concentrations (> 2M).  For many uni-cellular organisms, adaption to survival in these environments often occurs by the evolution of extremophilic proteins e.g. those that remain functional at low or high temperatures (psychrophilic and hyperthermophilic proteins).  In a collaboration with Lorna Dougan (Physics, Leeds), we have recently started to examine the biophysical and mechanical properties of a range of extremophilic proteins.

Publication: Single-molecule force spectroscopy identifies a small cold shock protein as being mechanically robust. Hoffmann et al. (2013) J Phys Chem B 117:1819−1826.

Exploring protein-ligand interactions.

In addition to characterising the behaviour of proteins under mechanical extension we have also started to measure the mechanical strength of the non-covalent interactions between proteins and their ligands. In particular we are interested in finding out how very strong interactions are broken apart on timescales fast enough to be biochemically useful. To do this we specifically immobilise one protein partner onto a surface and the other onto the tip of the AFM cantilever. This work has revealed that highly avid complexes can be rapidly broken apart by the application of small forces that are accessible to Nature.

Publication: A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein.   Farrance et al. (2013) PLoS Biol 11:e1001489.

Membrane protein folding.

Despite their ubiquity and importance as cellular gatekeepers, progress in understanding how membrane proteins fold into the narrow ensemble of structures required for their function is slow. In a collaboration with Professors Steve Baldwin and Sheena Radford we are examining the folding and insertion of the bacterial outer membrane protein PagP into liposomes and how periplasmic chaperones facilitate this process.

Publication: Malleability of the folding mechanism of the outer membrane protein PagP: parallel pathways and the effect of membrane elasticity.  Huysmans et al. J Mol Biol 416:453-463.

 

 

Detailed research programme                  Close ▲
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Associate Professor (Leeds) 2012 - present
BSc (Manchester) PhD (Manchester)


Postdoc (Manchester) 1997-1998
Postdoc (Leeds) 1998-2004
URF/Lecturer (Leeds) 2004-2012

Astbury 10.116
School of Molecular and Cellular Biology
0113 343 7821
d.j.brockwell@leeds.ac.uk
http://www.fbs.leeds.ac.uk/staff/profile.php?tag=Brockwell_D

Selected Publications

  1. Skp is a multivalent chaperone of outer-membrane proteins. Schiffrin, B., Calabrese, A., Devine, P., Harris, S., Ashcroft, A., Brockwell, D. and Radford, S. (2016) Nat Struct Mol Biol. 23:786––793.

  2. Rapid and robust polyprotein production facilitates single molecule mechanical characterization of of β-Barrel Assembly Machinery Polypeptide Transport Associated Domains.  Hoffmann, T., Tych, K., Crosskey, T., Schiffrin B., Dougan, L. and Brockwell, D. (2015) ACS Nano 9: 8811-8821.

  3. Extraction of accurate biomolecular parameters from single-molecule force spectroscopy experiments. Farrance, O., Paci, E., Radford, S. and Brockwell, D. (2015) ACS Nano 9: 1315-1324.

  4. A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein. Farrance, O., Hann, E., Kaminska, R., Derrington, S., Kleanthous, C., Radford, S. and Brockwell, D. (2013) PLoS Biol 11:e1001489