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Ortega Arroyo J.,Physical and Theoretical Chemistry Laboratory | Andrecka J.,Physical and Theoretical Chemistry Laboratory | Spillane K.M.,Physical and Theoretical Chemistry Laboratory | Billington N.,U.S. National Institutes of Health | And 3 more authors.
Nano Letters | Year: 2014

Optical detection of individual proteins requires fluorescent labeling. Cavity and plasmonic methodologies enhance single molecule signatures in the absence of any labels but have struggled to demonstrate routine and quantitative single protein detection. Here, we used interferometric scattering microscopy not only to detect but also to image and nanometrically track the motion of single myosin 5a heavy meromyosin molecules without the use of labels or any nanoscopic amplification. Together with the simple experimental arrangement, an intrinsic independence from strong electronic transition dipoles and a detection limit of <60 kDa, our approach paves the way toward nonresonant, label-free sensing and imaging of nanoscopic objects down to the single protein level. © 2014 American Chemical Society.

Degiacomi M.T.,Ecole Polytechnique Federale de Lausanne | Degiacomi M.T.,Physical and Theoretical Chemistry Laboratory | Dal Peraro M.,Ecole Polytechnique Federale de Lausanne
Structure | Year: 2013

Proteins often assemble in multimeric complexes to perform a specific biologic function. However, trapping these high-order conformations is difficult experimentally. Therefore, predicting how proteins assemble using in silico techniques can be of great help. The size of the associated conformational space and the fact that proteins are intrinsically flexible structures make this optimization problem extremely challenging. Nonetheless, known experimental spatial restraints can guide the search process, contributing to model biologically relevant states. We present here a swarm intelligence optimization protocol able to predict the arrangement of protein symmetric assemblies by exploiting a limited amount of experimental restraints and steric interactions. Importantly, within this scheme the native flexibility of each protein subunit is taken into account as extracted from molecular dynamics (MD) simulations. We show that this is a key ingredient for the prediction of biologically functional assemblies when, upon oligomerization, subunits explore activated states undergoing significant conformational changes. © 2013 Elsevier Ltd.

Wortham N.C.,University of Southampton | Martinez M.,University of Southampton | Gordiyenko Y.,Physical and Theoretical Chemistry Laboratory | Robinson C.V.,Physical and Theoretical Chemistry Laboratory | Proud C.G.,University of Southampton
FASEB Journal | Year: 2014

Eukaryotic initiation factor 2B (eIF2B) is the guanine nucleotide exchange factor for eIF2 and a critical regulator of protein synthesis, (e.g., as part of the integrated stress response). Certain mutations in the EIF2B genes cause leukoencephalopathy with vanishing white matter (VWM), an often serious neurological disorder. Comprising 5 subunits, α-ε (eIF2Bε being the catalytic one), eIF2B has always been considered an αβγ δε heteropentamer. We have analyzed the subunit interactions within mammalian eIF2B by using a combination of mass spectrometry and in vivo studies of overexpressed complexes to gain further insight into the subunit arrangement of the complex. Our data reveal that eIF2B is actually decameric, a dimer of eIF2B(βγδε) tetramers stabilized by 2 copies of eIF2Bα.We also demonstrate a pivotal role for eIF2Bδ in the formation of eIF2B(βγδε) tetramers. eIF2B(αβ γδε)2 decamers show greater binding to eIF2 than to eIF2B(βγδε) tetramers, which may underlie the increased activity of the former. We examined the levels of eIF2B subunits in a panel of different mouse tissues and identified different levels of eIF2B subunits, particularly eIF2Bα, which implies heterogeneity in the cellular proportions of eIF2B(αβγδε) and eIF2B(βγ δε) complexes, with important implications for the regulation of translation in individual cell types. © FASEB.

Shen H.-H.,Physical and Theoretical Chemistry Laboratory | Thomas R.K.,Physical and Theoretical Chemistry Laboratory | Taylor P.,Rutherford Appleton Laboratory
Langmuir | Year: 2010

We have investigated the formation of supported surfactin-phospholipid mixed bilayers using neutron reflectometry. Micellar mixtures of phospholipid (diphosphatidyl choline, DPPC), surfactin, and β-D-dodecyl maltoside were used to make the deposition. When the surfactin concentration is at its critical micelle concentration (CMC = 6 ×10- 6 M) in the bulk solution, there is no adsorption at all on the silica. When the surfactin concentration is lowered below the CMC, a mixed bilayer of surfactin and DPPC is formed. Since surfactin does not adsorb on silica from solutions of surfactin alone, this shows that there is a strong attraction between surfactin and DPPC. The variation of adsorbed amount, composition, and structure of the adsorbed layer are consistent with the attractive interaction between surfactin and DPPC and with their respective negative and positive affinities for the silica surface. Three phospholipid isotopic contrasts were measured and used to define the composition and structure of the surfactin-phospholipid bilayer. The maximum amount of surfactin in the bilayer reaches a mole fraction of about 0.2 and this is located in the outer leaflet of the bilayer within the headgroup and part of the adjacent chain region. © 2009 American Chemical Society.

Liebel M.,Physical and Theoretical Chemistry Laboratory | Schnedermann C.,Physical and Theoretical Chemistry Laboratory | Kukura P.,Physical and Theoretical Chemistry Laboratory
Physical Review Letters | Year: 2014

Coupling of nuclear and electronic degrees of freedom mediates energy flow in molecules after optical excitation. The associated coherent dynamics in polyatomic systems, however, remain experimentally unexplored. Here, we combined transient absorption spectroscopy with electronic population control to reveal nuclear wave packet dynamics during the S2→S1 internal conversion in β-carotene. We show that passage through a conical intersection is vibrationally coherent and thereby provides direct feedback on the role of different vibrational coordinates in the breakdown of the Born-Oppenheimer approximation. © 2014 American Physical Society.

Lu M.,Physical and Theoretical Chemistry Laboratory | Toghill K.E.,Physical and Theoretical Chemistry Laboratory | Compton R.G.,Physical and Theoretical Chemistry Laboratory
Electroanalysis | Year: 2011

We report the sensitive simultaneous detection of cadmium(II) and lead(II) using an unmodified edge plane pyrolytic graphite (EPPG) electrode via linear sweep anodic stripping voltammetry (LSASV). Simultaneous additions of the heavy metals gave two well separated stripping peaks observed over two linear ranges, 20 to 200μg L-1 and 2 to 20μg L-1. These gave detection limits of 0.3μg L-1 and 0.2μg L-1 for cadmium(II) and lead(II), respectively. The use of unmodified EPPG electrodes shows comparable performance to bismuth and mercury modified electrodes, and offers a simple and effective alternative to modified systems. © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Katelhon E.,Physical and Theoretical Chemistry Laboratory | Compton R.G.,Physical and Theoretical Chemistry Laboratory
Analyst | Year: 2014

The recent decade saw much interest in sensors based on nanoparticles. Such sensors typically employ sensing mechanisms that utilise the adsorption of analyte species on the nanoparticle surfaces, while adsorption induces changes in the physical properties of the nanoparticles. In this work, we introduce an analytical model for the rate of adsorption of analyte species on the nanoparticle surface. Expressions for the fractional surface coverage and the number of adsorbed molecules as a function of time are derived assuming spherical nanoparticles. Moreover, we provide values for common experimental conditions and show that for small nanoparticles (r < 10 nm) a surface coverages of 30% can be reached in less than 1 s at adsorbent concentrations as low as 50 nM. © the Partner Organisations 2014.

Lu M.,Physical and Theoretical Chemistry Laboratory | Compton R.G.,Physical and Theoretical Chemistry Laboratory
Electroanalysis | Year: 2013

A square wave voltammetric procedure for the determination of trace amounts of Fe(III) was developed at an unmodified edge plane pyrolytic graphite (EPPG) electrode and a screen printed electrode (SPE). This simple procedure was applied to real samples of commercially bottled mineral water. Sensitive results in the micromolar region could be achieved without modification of the electrode. Using the WHO guideline limits for the Fe(III) concentration in drinking water, recovery percentages at an EPPG gave 103% and 107%, and 98.6% and 95.0% at a SPE for the 5.36μM (0.3mgL-1) and 53.6μM (3.0mgL-1) additions of Fe(III), respectively. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Toghill K.E.,Physical and Theoretical Chemistry Laboratory | Compton R.G.,Physical and Theoretical Chemistry Laboratory
Electroanalysis | Year: 2010

Metal nanoparticle modified boron-doped diamond (BDD) electrodes have been used in the electroanalysis of a number of inorganic and organic analytes. This simple surface modification has enhanced the sensitivity and analytical ability of BDD effectively and consistently, as reported in a number of publications overviewed herein. In this review, a number of metal nanoparticle systems that have utilized BDD electrodes will be discussed. The low capacitance of BDD makes it an ideal substrate for sensitive dynamic electroanalytical experiments. The metal nanoparticle modification of BDD offers a simple yet effective approach to enhancing the electroanalytical ability of the electrode material. © 2010 Wiley-VCH Verlag GmbH&Co. KGaA, Weinheim.

Katelhon E.,Physical and Theoretical Chemistry Laboratory | Compton R.G.,Physical and Theoretical Chemistry Laboratory
Chemical Science | Year: 2014

We report the residence time of freely-diffusing, catalytically-active nanoparticles within a electron tunnelling distance of a surface. The role of near-wall hindered diffusion is paramount and leads to the new concept of "hydrodynamic adsorption". We give a comprehensive statistical analysis of the average impact times and derive expressions for a number values, crucial for the analysis of experimental data. Random walk simulations confirm the distribution of impact times with broad implications for nanochemistry. © the Partner Organisations 2014.

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