Peter Timmins

Publications (4)26.3 Total impact

• Source

  Available from: pnas.org

  Article: Absence of equilibrium cluster phase in concentrated lysozyme solutions.

  Anuj Shukla, Efstratios Mylonas, Emanuela Di Cola, Stephanie Finet, Peter Timmins, Theyencheri Narayanan, Dmitri I Svergun
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  ABSTRACT: In colloidal systems, the interplay between the short range attraction and long-range repulsion can lead to a low density associated state consisting of clusters of individual particles. Recently, such an equilibrium cluster phase was also reported for concentrated solutions of lysozyme at low ionic strength and close to the physiological pH. Stradner et al. [(2004) Equilibrium cluster formation in concentrated protein solutions and colloids. Nature 432:492-495] found that the position of the low-angle interference peak in small-angle x-ray and neutron scattering (SAXS and SANS) patterns from lysozyme solutions was essentially independent of the protein concentration and attributed these unexpected results to the presence of equilibrium clusters. This work prompted a series of experimental and theoretical investigations, but also revealed some inconsistencies. We have repeated these experiments following the protein preparation protocols of Stradner et al. using several batches of lysozyme and exploring a broad range of concentrations, temperature and other conditions. Our measurements were done in multiple experimental sessions at three different high-resolution SAXS and SANS instruments. The low-ionic-strength lysozyme solutions displayed a clear shift in peak positions with concentration, incompatible with the presence of the cluster phase but consistent with the system of repulsively interacting individual lysozyme molecules. Within the decoupling approximation, the experimental data can be fitted using an effective interparticle interaction potential involving short-range attraction and long-range repulsion.
  Proceedings of the National Academy of Sciences 05/2008; 105(13):5075-80. · 9.68 Impact Factor
• Source

  Available from: openrepository.com

  Article: X-ray and neutron small-angle scattering analysis of the complex formed by the Met receptor and the Listeria monocytogenes invasion protein InlB.

  Hartmut H Niemann, Maxim V Petoukhov, Michael Härtlein, Martine Moulin, Ermanno Gherardi, Peter Timmins, Dirk W Heinz, Dmitri I Svergun
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  ABSTRACT: The Listeria monocytogenes surface protein InlB binds to the extracellular domain of the human receptor tyrosine kinase Met, the product of the c-met proto-oncogene. InlB binding activates the Met receptor, leading to uptake of Listeria into normally nonphagocytic host cells. The N-terminal half of InlB (InlB(321)) is sufficient for Met binding and activation. The complex between this Met-binding domain of InlB and various constructs of the Met ectodomain was characterized by size exclusion chromatography and dynamic light scattering, and structural models were built using small-angle X-ray scattering and small-angle neutron scattering. Although most receptor tyrosine kinase ligands induce receptor dimerization, InlB(321) consistently binds the Met ectodomain with a 1:1 stoichiometry. A construct comprising the Sema and PSI domains of Met, although sufficient to bind the physiological Met ligand hepatocyte growth factor/scatter factor, does not form a complex with InlB(321) in solution, highlighting the importance of Met Ig domains for InlB binding. Small-angle X-ray scattering and small-angle neutron scattering measurements of ligand and receptor, both free and in complex, reveal an elongated shape for the receptor. The four Ig domains form a bent, rather than a fully extended, conformation, and InlB(321) binds to Sema and the first Ig domain of Met, in agreement with the recent crystal structure of a smaller Met fragment in complex with InlB(321). These results call into question whether receptor dimerization is the basic underlying event in InlB(321)-mediated Met activation and demonstrate differences in the mechanisms by which the physiological ligand hepatocyte growth factor/scatter factor and InlB(321) bind and activate the Met receptor.
  Journal of Molecular Biology 04/2008; 377(2):489-500. · 4.00 Impact Factor
• Article: A comparison of refined X-ray structures of hydrogenated and perdeuterated rat gammaE-crystallin in H2O and D2O.

  Jean Baptiste Artero, Michael Härtlein, Sean McSweeney, Peter Timmins
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  ABSTRACT: Rat gammaE-crystallin was overexpressed, purified under different labelling conditions and crystallized and X-ray data were collected at resolutions between 1.71 and 1.36 A. The structures were determined by molecular replacement. In these structures, the cd loop of the Greek-key motif 3, which is the major structural key motif of the two phase-transition groups of gamma-crystallins, presents a double conformation. The influence of the perdeuteration on the protein structure was determined by comparison of the atomic positions and temperature factors of the different models. The perdeuterated proteins have a similar structure to their hydrogenated counterparts, but partial or full deuteration may have some effect on the atomic B-factor values.
  Acta Crystallographica Section D Biological Crystallography 12/2005; 61(Pt 11):1541-9. · 12.62 Impact Factor
• Article: Reply to Stradner et al.: Equilibrium clusters are absent in concentrated lysozyme solutions

  Anuj Shukla, Efstratios Mylonas, Emanuela Di Cola, Stephanie Finet, Peter Timmins, Theyencheri Narayanan, Dmitri I Svergun