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Single molecule biophysics

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https://www.readbyqxmd.com/read/28087691/imaging-proteins-at-the-single-molecule-level
#1
Jean-Nicolas Longchamp, Stephan Rauschenbach, Sabine Abb, Conrad Escher, Tatiana Latychevskaia, Klaus Kern, Hans-Werner Fink
Imaging single proteins has been a long-standing ambition for advancing various fields in natural science, as for instance structural biology, biophysics, and molecular nanotechnology. In particular, revealing the distinct conformations of an individual protein is of utmost importance. Here, we show the imaging of individual proteins and protein complexes by low-energy electron holography. Samples of individual proteins and protein complexes on ultraclean freestanding graphene were prepared by soft-landing electrospray ion beam deposition, which allows chemical- and conformational-specific selection and gentle deposition...
January 13, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28076939/is-there-still-room-for-novelty-in-histochemical-papers
#2
Carlo Pellicciari
Histochemistry continues to be widely applied in biomedical research, being nowadays mostly addressed to detect and locate single molecules or molecular complexes inside cells and tissues, and to relate structural organization and function at the high resolution of the more advanced microscopical techniques. In the attempt to see whether histochemical novelties may be found in the recent literature, the articles published in the European Journal of Histochemistry in the period 2014-2016 have been reviewed. In the majority of the published papers, standardized methods have been preferred by scientists to make their results reliably comparable with the data in the literature, but  many papers (approximately one fourth of the published articles) described novel histochemical methods and procedures...
December 16, 2016: European Journal of Histochemistry: EJH
https://www.readbyqxmd.com/read/28069956/shuttling-along-dna-and-directed-processing-of-d-loops-by-recq-helicase-support-quality-control-of-homologous-recombination
#3
Gábor M Harami, Yeonee Seol, Junghoon In, Veronika Ferencziová, Máté Martina, Máté Gyimesi, Kata Sarlós, Zoltán J Kovács, Nikolett T Nagy, Yuze Sun, Tibor Vellai, Keir C Neuman, Mihály Kovács
Cells must continuously repair inevitable DNA damage while avoiding the deleterious consequences of imprecise repair. Distinction between legitimate and illegitimate repair processes is thought to be achieved in part through differential recognition and processing of specific noncanonical DNA structures, although the mechanistic basis of discrimination remains poorly defined. Here, we show that Escherichia coli RecQ, a central DNA recombination and repair enzyme, exhibits differential processing of DNA substrates based on their geometry and structure...
January 9, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28067976/single-nucleobase-identification-using-biophysical-signatures-from-nanoelectronic-quantum-tunneling
#4
Lee E Korshoj, Sepideh Afsari, Sajida Khan, Anushree Chatterjee, Prashant Nagpal
Nanoelectronic DNA sequencing can provide an important alternative to sequencing-by-synthesis by reducing sample preparation time, cost, and complexity as a high-throughput next-generation technique with accurate single-molecule identification. However, sample noise and signature overlap continue to prevent high-resolution and accurate sequencing results. Probing the molecular orbitals of chemically distinct DNA nucleobases offers a path for facile sequence identification, but molecular entropy (from nucleotide conformations) makes such identification difficult when relying only on the energies of lowest-unoccupied and highest-occupied molecular orbitals (LUMO and HOMO)...
January 9, 2017: Small
https://www.readbyqxmd.com/read/28062039/measuring-force-induced-dissociation-kinetics-of-protein-complexes-using-single-molecule-atomic-force-microscopy
#5
K Manibog, C F Yen, S Sivasankar
Proteins respond to mechanical force by undergoing conformational changes and altering the kinetics of their interactions. However, the biophysical relationship between mechanical force and the lifetime of protein complexes is not completely understood. In this chapter, we provide a step-by-step tutorial on characterizing the force-dependent regulation of protein interactions using in vitro and in vivo single-molecule force clamp measurements with an atomic force microscope (AFM). While we focus on the force-induced dissociation of E-cadherins, a critical cell-cell adhesion protein, the approaches described here can be readily adapted to study other protein complexes...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28062035/single-stranded-dna-curtains-for-studying-homologous-recombination
#6
C J Ma, J B Steinfeld, E C Greene
Homologous recombination is an important pathway involved in the repair of double-stranded DNA breaks. Genetic studies form the foundation of our knowledge on homologous recombination. Significant progress has also been made toward understanding the biochemical and biophysical properties of the proteins, complexes, and reaction intermediates involved in this essential DNA repair pathway. However, heterogeneous or transient recombination intermediates remain extremely difficult to assess through traditional ensemble methods, leaving an incomplete mechanistic picture of many steps that take place during homologous recombination...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28059871/modified-nanoantibodies-increase-sensitivity-in-avidin-biotin-immunohistochemistry
#7
Anthony Wong, Chelsea Sykora, Lewis Rogers, Jennifer Higginbotham, Jiwu Wang
Revealing the spatial arrangement of molecules within a tissue through immunohistochemistry (IHC) is an invaluable tool in biomedical research and clinical diagnostics. Choosing both the appropriate antibody and amplification system is paramount to the pathologic interpretation of the tissue at hand. The use of single domain VHH nanoantibodies (nAbs) promise more robust and consistent results in IHC, but are rarely used as an alternative to conventional immunoglobulin G (IgG) antibodies. nAbs are originally obtained from llamas and are the smallest antigen-binding fragments available...
January 4, 2017: Applied Immunohistochemistry & Molecular Morphology: AIMM
https://www.readbyqxmd.com/read/28032989/how-does-your-protein-fold-elucidating-the-apomyoglobin-folding-pathway
#8
H Jane Dyson, Peter E Wright
Although each type of protein fold and in some cases individual proteins within a fold classification can have very different mechanisms of folding, the underlying biophysical and biochemical principles that operate to cause a linear polypeptide chain to fold into a globular structure must be the same. In an aqueous solution, the protein takes up the thermodynamically most stable structure, but the pathway along which the polypeptide proceeds in order to reach that structure is a function of the amino acid sequence, which must be the final determining factor, not only in shaping the final folded structure, but in dictating the folding pathway...
December 29, 2016: Accounts of Chemical Research
https://www.readbyqxmd.com/read/28018237/multidimensional-liquid-chromatography-coupled-with-tandem-mass-spectrometry-for-identification-of-bioactive-fatty-acyl-derivatives
#9
Erin B Divito, Kristin M Kroniser, Michael Cascio
Recognition of the contributions of lipids to cellular physiology, both as structural components of the membrane and as modulatory ligands for membrane proteins, has increased in recent years with the development of the biophysical and biochemical tools to examine these effects. Their modulatory roles in ion channels and transporters function have been extensively characterized, with the molecular mechanisms of these activities being the subject of intense scrutiny. The physiological significance of lipids in biochemistry is expanding as numerous fatty acyls are discovered to possess signaling properties...
2016: Frontiers in Physiology
https://www.readbyqxmd.com/read/27982056/microstructure-imaging-of-crossing-mix-white-matter-fibers-from-diffusion-mri
#10
Hamza Farooq, Junqian Xu, Jung Who Nam, Daniel F Keefe, Essa Yacoub, Tryphon Georgiou, Christophe Lenglet
Diffusion MRI (dMRI) reveals microstructural features of the brain white matter by quantifying the anisotropic diffusion of water molecules within axonal bundles. Yet, identifying features such as axonal orientation dispersion, density, diameter, etc., in complex white matter fiber configurations (e.g. crossings) has proved challenging. Besides optimized data acquisition and advanced biophysical models, computational procedures to fit such models to the data are critical. However, these procedures have been largely overlooked by the dMRI microstructure community and new, more versatile, approaches are needed to solve complex biophysical model fitting problems...
December 16, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27966899/extracellular-osmotic-stress-reduces-the-vesicle-size-while-keeping-a-constant-neurotransmitter-concentration
#11
Hoda Fathali, Johan Dunevall, Soodabeh Majdi, Ann-Sofie Cans
Secretory cells respond to hypertonic stress by cell shrinking, which causes a reduction in exocytosis activity and the amount of signaling molecules released from single exocytosis events. These changes in exocytosis have been suggested to result from alterations in biophysical properties of cell cytoplasm and plasma membrane, based on the assumption that osmotic stress does not affect the secretory vesicle content and size prior to exocytosis. To further investigate whether vesicles in secretory cells are affected by the osmolality of the extracellular environment, we used intracellular electrochemical cytometry together with transmission electron microscopy imaging to quantify and determine the catecholamine concentration of dense core vesicles in situ before and after cell exposure to osmotic stress...
January 4, 2017: ACS Chemical Neuroscience
https://www.readbyqxmd.com/read/27960258/mechanism-of-the-spontaneous-and-directional-membrane-insertion-of-a-2-transmembrane-ion-channel
#12
Steffen Altrichter, Maximilian Haase, Belinda Loh, Andreas Kuhn, Sebastian Leptihn
Protein insertion into membranes is a process occurring in every cell and every cellular compartment. Yet, many thermodynamic aspects of this fundamental biophysical process are not well understood. We investigated physicochemical parameters that influence protein insertion using the model protein KcsA, a 2-transmembrane ion channel. To understand what drives insertion and to identify individual steps of protein integration into a highly apolar environment, we investigated the contribution of electrostatic interactions and lipid composition on protein insertion on a single molecule level...
December 21, 2016: ACS Chemical Biology
https://www.readbyqxmd.com/read/27934066/nanopores-and-nanochannels-from-gene-sequencing-to-genome-mapping
#13
Stefan Howorka, Zuzanna Siwy
DNA strands can be analyzed at the single-molecule level by isolating them inside nanoscale holes. The strategy is used for the label-free and portable sequencing with nanopores. Nanochannels can also be applied to map genomes with high resolution, as shown by Jeffet et al. in this issue of ACS Nano. Here, we compare the two strategies in terms of biophysical similarities and differences and describe that both are complementary and can improve the DNA analysis for genomic research and diagnostics.
November 22, 2016: ACS Nano
https://www.readbyqxmd.com/read/27933808/accurate-modeling-of-scaffold-hopping-transformations-in-drug-discovery
#14
Lingle Wang, Yuqing Deng, Yujie Wu, Byungchan Kim, David N LeBard, Dan Wandschneider, Mike Beachy, Richard A Friesner, Robert Abel
The accurate prediction of protein-ligand binding free energies remains a significant challenge of central importance in computational biophysics and structure-based drug design. Multiple recent advances including the development of greatly improved protein and ligand molecular mechanics force fields, more efficient enhanced sampling methods, and low-cost powerful GPU computing clusters have enabled accurate and reliable predictions of relative protein-ligand binding free energies through the free energy perturbation (FEP) methods...
December 9, 2016: Journal of Chemical Theory and Computation
https://www.readbyqxmd.com/read/27870840/single-nanotube-tracking-reveals-the-nanoscale-organization-of-the-extracellular-space-in-the-live-brain
#15
Antoine G Godin, Juan A Varela, Zhenghong Gao, Noémie Danné, Julien P Dupuis, Brahim Lounis, Laurent Groc, Laurent Cognet
The brain is a dynamic structure with the extracellular space (ECS) taking up almost a quarter of its volume. Signalling molecules, neurotransmitters and nutrients transit via the ECS, which constitutes a key microenvironment for cellular communication and the clearance of toxic metabolites. The spatial organization of the ECS varies during sleep, development and aging and is probably altered in neuropsychiatric and degenerative diseases, as inferred from electron microscopy and macroscopic biophysical investigations...
November 21, 2016: Nature Nanotechnology
https://www.readbyqxmd.com/read/27844425/beyond-the-hookean-spring-model-direct-measurement-of-optical-forces-through-light-momentum-changes
#16
Arnau Farré, Ferran Marsà, Mario Montes-Usategui
The ability to measure forces in the range of 0.1-100 pN is a key feature of optical tweezers used for biophysical and cell biological studies. Analysis of the interactions between biomolecules and the forces that biomolecular motors generate at the single-molecule level has provided valuable insights in the molecular mechanisms that govern key cellular functions such as gene expression and the long-distance transport of organelles. Methods for determining the minute forces that biomolecular motors generate exhibit notable constraints that limit their application for studies other than the well-controlled in vitro experiments (although recent advances have been made that permit more quantitative optical tweezers studies insight living cells)...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/27833562/predicting-effects-of-tropomyosin-mutations-on-cardiac-muscle-contraction-through-myofilament-modeling
#17
Lorenzo R Sewanan, Jeffrey R Moore, William Lehman, Stuart G Campbell
Point mutations to the human gene TPM1 have been implicated in the development of both hypertrophic and dilated cardiomyopathies. Such observations have led to studies investigating the link between single residue changes and the biophysical behavior of the tropomyosin molecule. However, the degree to which these molecular perturbations explain the performance of intact sarcomeres containing mutant tropomyosin remains uncertain. Here, we present a modeling approach that integrates various aspects of tropomyosin's molecular properties into a cohesive paradigm representing their impact on muscle function...
2016: Frontiers in Physiology
https://www.readbyqxmd.com/read/27806271/functional-analysis-of-orai1-concatemers-supports-a-hexameric-stoichiometry-for-the-crac-channel
#18
Michelle Yen, Ludmila A Lokteva, Richard S Lewis
Store-operated Ca(2+) entry occurs through the binding of the endoplasmic reticulum (ER) Ca(2+) sensor STIM1 to Orai1, the pore-forming subunit of the Ca(2+) release-activated Ca(2+) (CRAC) channel. Although the essential steps leading to channel opening have been described, fundamental questions remain, including the functional stoichiometry of the CRAC channel. The crystal structure of Drosophila Orai indicates a hexameric stoichiometry, while studies of linked Orai1 concatemers and single-molecule photobleaching suggest that channels assemble as tetramers...
November 1, 2016: Biophysical Journal
https://www.readbyqxmd.com/read/27784615/quantitative-analysis-of-the-role-of-fiber-length-on-phagocytosis-and-inflammatory-response-by-alveolar-macrophages
#19
Trudy Padmore, Carahline Stark, Leonid A Turkevich, Julie A Champion
BACKGROUND: In the lung, macrophages attempt to engulf inhaled high aspect ratio pathogenic materials, secreting inflammatory molecules in the process. The inability of macrophages to remove these materials leads to chronic inflammation and disease. How the biophysical and biochemical mechanisms of these effects are influenced by fiber length remains undetermined. This study evaluates the role of fiber length on phagocytosis and molecular inflammatory responses to non-cytotoxic fibers, enabling development of quantitative length-based models...
February 2017: Biochimica et Biophysica Acta
https://www.readbyqxmd.com/read/27783939/camp-signals-in-drosophila-motor-neurons-are-confined-to-single-synaptic-boutons
#20
Isabella Maiellaro, Martin J Lohse, Robert J Kittel, Davide Calebiro
The second messenger cyclic AMP (cAMP) plays an important role in synaptic plasticity. Although there is evidence for local control of synaptic transmission and plasticity, it is less clear whether a similar spatial confinement of cAMP signaling exists. Here, we suggest a possible biophysical basis for the site-specific regulation of synaptic plasticity by cAMP, a highly diffusible small molecule that transforms the physiology of synapses in a local and specific manner. By exploiting the octopaminergic system of Drosophila, which mediates structural synaptic plasticity via a cAMP-dependent pathway, we demonstrate the existence of local cAMP signaling compartments of micrometer dimensions within single motor neurons...
October 25, 2016: Cell Reports
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