Department of Biochemistry University of Oxford Department of Biochemistry
University of Oxford
South Parks Road
Oxford OX1 3QU

Tel: +44 (0)1865 613200
Fax: +44 (0)1865 613201
Image showing the global movement of lipids in a model planar membrane
Matthieu Chavent, Sansom lab
Anaphase bridges in fission yeast cells
Whitby lab
Lactose permease represented using bending cylinders in Bendix software
Caroline Dahl, Sansom lab
Epithelial cells in C. elegans showing a seam cell that failed to undergo cytokinesis
Serena Ding, Woollard lab
Collage of Drosophila third instar larva optic lobe
Lu Yang, Davis lab
First year Biochemistry students at a practical class
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Colin Kleanthous
Protein-protein interactions in the Gram-negative bacterial cell envelope

Co-workers: Nick Housden, Renata Kaminska, Joanna Szczepaniak, Sandip Kumar, Melissa Webby, Ruth Cohen-Khait, Gideon Mamou (EMBO LTF), Soumik Basu (RS Newton International fellow), Connor Sharp, Nathalie Reichmann, Marie-Louise Francis, Hannah Behrens, Jonathan Goult, Patrick Inns, Iva Atanaskovic

In my laboratory, we aim to understand how protein-protein interactions (PPIs) in bacteria underpin organisation of the bacterial outer membrane and Tol-Pal signalling across the bacterial cell envelope and how these signals are subverted by antibacterial proteins (bacteriocins) to catalyse their import into the cell. More broadly, we are interested in developing bacteriocins as antibiotics to target Gram-negative pathogens. We adopt a multidisciplinary approach in dissecting biological function that incorporates protein chemistry and engineering, molecular biophysics, structural biology and in vivo imaging. Our work is supported by the Wellcome Trust, ERC, BBSRC and MRC.

Bacteriocin translocation – Bacteriocins are species-specific protein antibiotics that parasitize a variety of outer membrane and periplasmic proteins in Gram-negative bacteria. Much of our work has focused on the entry mechanism of colicins, which target Escherichia coli , but we have also begun studies on pyocins, which target Pseudomonas aeruginosa , klebicins, which target Klebsiella pneumoniae , and marcescins, which target Serratia marcescens . These toxins serve as important agents of competition within microbial communities. We study both nuclease (DNases, rRNases and tRNases) and pore-forming bacteriocins, which use their network of PPIs within the cell envelope to establish a translocon complex that delivers a toxic domain into the cell. Hence, bacteriocin translocation represents a highly simplified model for cellular protein import.

Developing bacteriocins as protein antibiotics– The rise of multidrug resistant bacteria coupled with the lack of new classes of antibiotics in over 30 years means there is a pressing and urgent need for new antibiotics especially for molecules that target pathogenic Gram-negative bacteria.

In collaboration with groups in Oxford, Glasgow, the Sanger Institute and UCL, we are exploiting the species selectivity and potency of bacteriocins as antimicrobials for Pseudomonas aeruginosa and Klebsiella pneumoniae, which are major causes of opportunistic and hospital-acquired infections world-wide.

Supramolecular assembly of outer membrane proteins– The asymmetric bilayer of the outer membrane contains outer membrane proteins (OMPs) and lipoproteins. Using colicins as OMP-specific probes, we recently discovered that OMPs cluster to form ‘OMP islands’ in the membrane and that this clustering behaviour underpins their turnover. We also demonstrated that the clustering behaviour of OMPs could be replicated in vitro using supported bilayers. Through the creation of a variety of OMP-specific labels we are dissecting the organisation of OMP islands and investigating the ramifications of these micro-domains to bacterial physiology. For example, we demonstrated recently that the inherent organisation of outer membrane proteins into islands becomes imprinted on inner membrane proteins when they are connected by an energized protein bridge.

The Tol-Pal assembly – Tol-Pal is a little understood complex that is required for the stable maintenance of the Gram-negative outer membrane and which is recruited to the septation apparatus during cell division. We are trying to uncover the native function of the Tol-Pal assembly, how and why it is coupled to the proton-motive force across the inner membrane and the mechanism by which bacteriocins subvert the assembly to initiate import across the outer membrane.

Selection of Recent Publications

  1. Rassam, P., Long, K.R., Kaminsak, R., Williams, D.J., Papadakos, G., Baumann, C.G.* & Kleanthous, C.* (2018) Intermembrane crosstalk drives inner membrane protein organization in Escherichia coli. Nat. Commun. 9, 1082.
  2. White, P., Joshi, A., Rassam, P., Housden, N.G., Kaminska, R., Goult, J.D., Redfield, C., McCaughey, L.C., Walker, D., Mohammed, S. & Kleanthous, C. (2017) Exploitation of an iron transporter for bacterial protein antibiotic import. Proc. Natl. Acad. Sci. USA 114, 12051-12056.
  3. Rajasekar, K.V., Zdanowski, K., Yan, J., Hopper, J.T.S., Francis, M.L.R., Seepersad, C., Sharp, C., Pecqueur, L., Werner, J.M., Robinson, C.V., Mohammed, S., Potts, J. & Kleanthous, C. (2016) The anti-sigma factor RsrA responds to oxidative stress by reburying its hydrophobic core. Nat. Commun. 7, 12194.
  4. Rassam, P., Copeland, N.A., Birkholz, O., Tóth, C., Chavent, M., Duncan, A.L., Cross, S.J., Housden, N.G., Kaminska, R., Seger, U., Quinn, D.M., Garrod, T.J., Sansom, M.S.P., Piehler, J., Baumann, C.G.* & Kleanthous, C.* (2015) Supramolecular assemblies underpin turnover of outer membrane proteins in bacteria. Nature 523, 333-336.
    Subject of Trends in Microbiology Spotlight review (2015) 23, 452
  5. Housden, N.G., Hopper, J.T.S., Lukoyanova, N., Rodriguez-Larrea, D., Wojdyla, J.A., Klein, A., Kaminska, R., Bayley, H., Saibil, H.R., Robinson, C.V. & Kleanthous, C. (2013) Intrinsically disordered protein threads through the bacterial outer membrane porin OmpF. Science 340, 1570-1574.
    Selected as a Leading Edge article on Intrinsically Disordered Proteins (2013) Cell 154, 473
More Publications...

Research Images


Figure 1. Directed epitope delivery, a novel transmembrane signalling mechanism exploited by colicins. Colicin E9 (ColE9) houses two OmpF-binding sites within its 83-residue intrinsically unstructured translocation domain (IUTD). The colicin manages to capture TolB on the other side of the membrane by threading its IUTD through two pores of an OmpF trimer, one going into the cell the other coming back out again. In this way TolB is held in a defined orientation relative to the TolQRA inner membrane complex contact with which triggers entry of the toxin. See ref. 5 for further details.

Figure 2. OMPs co-localize in supramolecular assemblies in the outer membrane of E. coliFigure adapted from ref. 1. Left-hand panel, engineering colicins E9 and Ia as specific, high affinity labels of the OMPs BtuB and Cir. BtuB is the transporter for vitamin B12 while Cir is a siderophore transporter. The two colicins were inactivated by internal disulphide bonds prior to labelling with fluorophores. Right-hand panel, E. coli OMP islands imaged by total internal reflection fluorescence microscopy and showing the co-localization of BtuB (labelled with ColE9-AF488, green) and Cir (labelled with ColIa-TMR, red). Scale bar is 1 mm. See ref. 4 for details.

Figure 3. BtuB-bound ColE9 induces clustering in the inner membrane protein TolA. a , 3D-SIM images of E. coli cells expressing GFP-TolA showing how formation of a transenvelope bridge by ColE9 (as in Figure 1) induces clustering of TolA in the inner membrane (IM). b, 2D-SIM z-slice showing significant co-clustering ( yellow fluorescence) of GFP-TolA and ColE9AF594 in the IM and OM, respectively. c, TIRFM data showing co-localization of GFP-TolA clusters in the IM and OMP-bound ColE9 TMR islands (yellow fluorescence). See ref. 1 for details.



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