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Single molecule biophysics

Raju Regmi, Pamina Martina Winkler, Valentin Flauraud, Kyra Borgman, Carlo Manzo, Juergen Brugger, Herve Rigneault, Jerome Wenger, Maria F Garcia-Parajo
Optical nano-antennas can efficiently confine light into nanoscopic hotspots enabling single-molecule detection sensitivity at biological relevant conditions. This innovative approach to breach the diffraction limit offers a versatile platform to investigate the dynamics of individual biomolecules in living cell membranes and their partitioning into cholesterol-dependent lipid nanodomains. Here, we present optical nano-antenna arrays with accessible surface hotspots to study the characteristic diffusion dynamics of phosphoethanolamine (PE) and sphingomyelin (SM) in the plasma membrane of living cells at the nanoscale...
September 19, 2017: Nano Letters
Jen-Wei Liu, Chih-Wen Cheng, Yu-Feng Lin, Shao-Yu Chen, Jenn-Kang Hwang, Shih-Chung Yen
BACKGROUND: Functional and biophysical constraints can cause different levels of sequence conservation in proteins. Previously, structural properties, e.g., relative solvent accessibility (RSA) and packing density of the weighted contact number (WCN), have been found to be related to protein sequence conservation (CS). The Voronoi volume has recently been recognized as a new structural property of the local protein structural environment reflecting CS. However, for surface residues, it is sensitive to water molecules surrounding the protein structure...
September 11, 2017: Biochimica et Biophysica Acta
Seoyoung Son, George J Moroney, Peter J Butler
Integrin-mediated adhesion is a central feature of cellular adhesion, locomotion, and endothelial cell mechanobiology. Although integrins are known to be transmembrane proteins, little is known about the role of membrane biophysics and dynamics in integrin adhesion. We treated human aortic endothelial cells with exogenous amphiphiles, shown previously in model membranes, and computationally, to affect bilayer thickness and lipid phase separation, and subsequently measured single-integrin-molecule adhesion kinetics using an optical trap, and diffusion using fluorescence correlation spectroscopy...
September 5, 2017: Biophysical Journal
Helen Miller, Zhaokun Zhou, Jack Shepherd, Adam Wollman, Mark Leake
Single-molecule biophysics has transformed our understanding of biology, but also of the physics of life. More exotic than simple soft matter, biomatter lives far from thermal equilibrium, covering multiple lengths from the nanoscale of single molecules to up several orders of magnitude to higher in cells, tissues and organisms. Biomolecules are often characterized by underlying instability: multiple metastable free energy states exist, separated by levels of just a few multiples of the thermal energy scale kBT, where kB is the Boltzmann constant and T absolute temperature, implying complex inter-conversion kinetics in the relatively hot, wet environment of active biological matter...
September 4, 2017: Reports on Progress in Physics
(no author information available yet)
Ana Garcia-Saez graduated with a master's degree in biochemistry from the University of Valencia, Spain, before completing a PhD in the laboratory of Jesus Salgado in 2005. Afterwards Ana joined the research group of Petra Schwille at the Dresden University of Technology, Germany. In 2010, she started her own group at BioQuant, the Center for Quantitative Analysis of Molecular and Cellular Biosystems, at Heidelberg University with funding from the Max Planck Institute for Intelligent Systems and the German Cancer Research Centre (DKFZ)...
September 1, 2017: Journal of Cell Science
Paolo Cadinu, Binoy Paulose Nadappuram, Dominic Lee, Jasmine Y Y Sze, Giulia Campolo, Yanjun Zhang, Andrew Shevchuk, Sylvain Ladame, Tim Albrecht, Yuri E Korchev, Aleksandar P Ivanov, Joshua B Edel
There is a growing realization, especially within the diagnostic and therapeutic community, that the amount of information enclosed in a single molecule can not only enable a better understanding of biophysical pathways, but can also offer exceptional value for early stage biomarker detection of disease onset. To this end, numerous single molecule strategies have been proposed and in terms of label-free routes, nanopore sensing has emerged as one of the most promising methods. However, being able to finely control molecular transport in terms of transport rate, resolution, and signal-to-noise ratio (SNR) is essential to take full advantage of the technology benefits...
September 1, 2017: Nano Letters
Tuanlao Wang, Liangcheng Li, Wanjin Hong
SNAREs (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptors) are the core machinery mediating membrane fusion. In this review, we provide an update on the recent progress on SNAREs regulating membrane fusion events, especially, the more detailed fusion processes dissected by well developed biophysical methods and in vitro single molecule analysis approaches. We also briefly summarize the relevant research from Chinese laboratories and highlight the significant contributions on our understanding of SNARE mediated membrane trafficking from scientists in China...
August 31, 2017: Traffic
Ekaterina L Grishchuk
The main physiological function of mitotic kinetochores is to provide durable attachment to spindle microtubules, which segregate chromosomes in order to partition them equally between the two daughter cells. Numerous kinetochore components that can bind directly to microtubules have been identified, including ATP-dependent motors and various microtubule-associated proteins with no motor activity. A major challenge facing the field is to explain chromosome motions based on the biochemical and structural properties of these individual kinetochore components and their assemblies...
2017: Progress in Molecular and Subcellular Biology
Phoebe S Tsoi, Kyoung-Jae Choi, Paul G Leonard, Antons Sizovs, Mahdi Muhammad Moosa, Kevin R MacKenzie, Josephine C Ferreon, Allan Chris M Ferreon
Transactivation response element (TAR) DNA-binding protein 43 (TDP-43) protein misfolding is implicated in several neurodegenerative diseases characterized by aggregated protein inclusions. Misfolding is believed to be mediated by both the N- and C-terminus of TDP-43; however, the mechanistic basis of the contribution of individual domains in the process remained elusive. Here, using single-molecule fluorescence and ensemble biophysical techniques, and a wide range of pH and temperature conditions, we show that TDP-43NTD is thermodynamically stable, well-folded and undergoes reversible oligomerization...
August 21, 2017: Angewandte Chemie
Caroline Senac, Wladimir Urbach, Erol Kurtisovski, Philippe H Hünenberger, Bruno A C Horta, Nicolas Taulier, Patrick F J Fuchs
Polyoxyethylene glycol alkyl ether amphiphiles (CiEj) are important nonionic surfactants, often used for biophysical and membrane protein studies. In this work, we extensively test the GROMOS-compatible 2016H66 force field in molecular dynamics simulations involving the lamellar phase of a series of CiEj surfactants, namely C12E2, C12E3, C12E4, C12E5, and C14E4. The simulations reproduce qualitatively well the monitored structural properties and their experimental trends along the surfactant series, although some discrepancies remain, in particular in terms of the area per surfactant, the equilibrium phase of C12E5, and the order parameters of C12E3, C12E4, and C12E5...
September 20, 2017: Langmuir: the ACS Journal of Surfaces and Colloids
Tze-Yun Huang, Chung-Ke Chang, Ya-Fen Kao, Chih-Hao Chin, Cheng-Wei Ni, Hao-Yi Hsu, Nien-Jen Hu, Li-Ching Hsieh, Shan-Ho Chou, I-Ren Lee, Ming-Hon Hou
Repetitive DNA sequences are ubiquitous in life, and changes in the number of repeats often have various physiological and pathological implications. DNA repeats are capable of interchanging between different noncanonical and canonical conformations in a dynamic fashion, causing configurational slippage that often leads to repeat expansion associated with neurological diseases. In this report, we used single-molecule spectroscopy together with biophysical analyses to demonstrate the parity-dependent hairpin structural polymorphism of TGGAA repeat DNA...
August 21, 2017: Proceedings of the National Academy of Sciences of the United States of America
Carolina Sanchez, Lena Voith-von-Voithenberg, Lisa Warner, Don C Lamb, Michael Sattler
Fluorescence-based techniques are widely used to study biomolecular conformations, intra- and intermolecular interactions, and conformational dynamics of macromolecules. Especially for fluorescent-based single-molecule experiments, the choice of the fluorophore and labeling position are highly important. Here, we have studied the biophysical and structural effects that are associated with the conjugation of fluorophores to cysteines in the splicing factor U2AF65 by using single pair Förster resonance energy transfer (FRET) and nuclear magnetic resonance (NMR) spectroscopy...
August 10, 2017: Chemistry: a European Journal
Ryota Iino, Tatsuya Iida, Akihiko Nakamura, Ei-Ichiro Saita, Huijuan You, Yasushi Sako
Biological molecular machines support various activities and behaviors of cells, such as energy production, signal transduction, growth, differentiation, and migration. Scope of Review We provide an overview of single-molecule imaging methods involving both small and large probes used to monitor the dynamic motions of molecular machines in vitro (purified proteins) and in living cells, and single-molecule manipulation methods used to measure the forces, mechanical properties and responses of biomolecules. We also introduce several examples of single-molecule analysis, focusing primarily on motor proteins and signal transduction systems...
August 5, 2017: Biochimica et Biophysica Acta
Michael Staniforth, Wen-Dong Quan, Tolga N V Karsili, Lewis A Baker, Rachel K O'Reilly, Vasilios G Stavros
Continuous advancements in biophysics and medicine at the molecular level make the requirements to image structure-function processes in living cells ever more acute. While fluorophores such as the green fluorescent protein have proven instrumental toward such efforts, the advent of nondiffraction limited microscopy limits the utility of such fluorescent tags. Monoaminomaleimides are small, single molecule fluorophores that have been shown to possess stark variations in their emission spectra in different solvent environments, making them a potentially powerful tool for a myriad of applications...
August 23, 2017: Journal of Physical Chemistry. A
Anthony Fernandez, Markville Bautista, Ramunas Stanciauskas, Taerin Chung, Fabien Pinaud
Patterning cells on microcontact-printed substrates is a powerful approach to control cell morphology and introduce specific mechanical cues on a cell's molecular organization. Although global changes in cellular architectures caused by micropatterns can easily be probed with diffraction-limited optical microscopy, studying molecular reorganizations at the nanoscale demands micropatterned substrates that accommodate the optical requirements of single molecule microscopy techniques. Here, we developed a simple micropatterning strategy that provides control of cellular architectures and is optimized for nanometer accuracy single molecule tracking and three-dimensional super-resolution imaging of plasma and nuclear membrane proteins in cells...
August 9, 2017: ACS Applied Materials & Interfaces
A Gilardi, S P Bhamidimarri, M Brönstrup, U Bilitewski, R K R Marreddy, K M Pos, L Benier, P Gribbon, M Winterhalter, B Windshügel
BACKGROUND: The tripartite efflux pump AcrAB-TolC in E. coli is involved in drug resistance by transporting antibiotics out of the cell. The outer membrane protein TolC can be blocked by various cations, including hexaamminecobalt, thereby TolC represents a potential target for reducing antimicrobial resistance as its blockage may improve efficacy of antibiotics. METHODS: We utilized single channel electrophysiology measurements for studying TolC conductance in the absence and presence of the known TolC blocker hexaamminecobalt...
July 23, 2017: Biochimica et Biophysica Acta
Pawel R Laskowski, Moritz Pfreundschuh, Mirko Stauffer, Zöhre Ucurum, Dimitrios Fotiadis, Daniel J Müller
To understand how membrane proteins function requires characterizing their structure, assembly, and inter- and intramolecular interactions in physiologically relevant conditions. Conventionally, such multiparametric insight is revealed by applying different biophysical methods. Here we introduce the combination of confocal microscopy, force-distance curve-based (FD-based) atomic force microscopy (AFM), and single-molecule force spectroscopy (SMFS) for the identification of native membranes and the subsequent multiparametric analysis of their membrane proteins...
August 22, 2017: ACS Nano
Emmanuel Derivery, Eline Bartolami, Stefan Matile, Marcos Gonzalez-Gaitan
Quantum dots (QDs) are extremely bright, photostable, nanometer particles broadly used to investigate single molecule dynamics in vitro. However, the use of QDs in vivo to investigate single molecule dynamics is impaired by the absence of an efficient way to chemically deliver them into the cytosol of cells. Indeed, current methods (using cell-penetrating peptides for instance) provide very low yields: QDs stay at the plasma membrane or are trapped in endosomes. Here, we introduce a technology based on cell-penetrating poly(disulfide)s that solves this problem: we deliver about 70 QDs per cell, and 90% appear to freely diffuse in the cytosol...
August 2, 2017: Journal of the American Chemical Society
Takayuki Uchihashi, Simon Scheuring
BACKGROUND: Many biological processes in a living cell are consequences of sequential and hierarchical dynamic events of biological macromolecules such as molecular interactions and conformational changes. Hence, knowledge of structures, assembly and dynamics of proteins is the foundation for understanding how biological molecules work. Among several techniques to analyze dynamics of proteins, high-speed atomic force microscopy (HS-AFM) is unique to provide direct information about both structure and dynamics of single proteins at work...
July 14, 2017: Biochimica et Biophysica Acta
Javier Cervera, Salvador Meseguer, Salvador Mafe
We have studied theoretically the microRNA (miRNA) intercellular transfer through voltage-gated gap junctions in terms of a biophysically grounded system of coupled differential equations. Instead of modeling a specific system, we use a general approach describing the interplay between the genetic mechanisms and the single-cell electric potentials. The dynamics of the multicellular ensemble are simulated under different conditions including spatially inhomogeneous transcription rates and local intercellular transfer of miRNAs...
August 2, 2017: Journal of Physical Chemistry. B
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