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Single molecule biophysics

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https://www.readbyqxmd.com/read/27870840/single-nanotube-tracking-reveals-the-nanoscale-organization-of-the-extracellular-space-in-the-live-brain
#1
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
#2
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
#3
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
#4
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
#5
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...
October 23, 2016: Biochimica et Biophysica Acta
https://www.readbyqxmd.com/read/27783939/camp-signals-in-drosophila-motor-neurons-are-confined-to-single-synaptic-boutons
#6
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
https://www.readbyqxmd.com/read/27749866/visualizing-the-molecular-mode-of-motion-from-a-correlative-analysis-of-localization-microscopy-datasets
#7
Carmine Di Rienzo, Paolo Annibale
The determination of the mode and rapidity of motion of individual molecules within a biological sample is becoming a more and more common analysis in biophysical investigations. Single molecule tracking (SMT) techniques allow reconstructing the trajectories of individual molecules within a movie, provided that the position from one frame to the other can be correctly linked. The outcomes, however, appear to depend on the specific method used, and most techniques display a limitation to capture fast modes of motion in a crowded environment...
October 1, 2016: Optics Letters
https://www.readbyqxmd.com/read/27744697/single-molecule-fret-studies-of-hybridization-mechanism-during-the-noncovalent-adsorption-and-desorption-of-dna-on-graphene-oxide
#8
Tapas Paul, Subhas Chandra Bera, Nidhi Agnihotri, Padmaja P Mishra
A remarkable observation about adsorption and desorption mechanism of single-stranded oligo nucleotides and hybridization of double stranded DNA (ds-DNA) on graphene oxide (GO) surface has been monitored using ensemble and single molecule fluorescence methods. The probe and target DNA, both labeled individually with fluorescence resonance energy transfer (FRET) pair and having similar adsorption affinity towards the GO surface are used to provide the detailed insight into the hybridization mechanism. The single molecule-FRET (sm-FRET) results reveals the DNA hybridization mechanism to happen "in situ", i...
October 17, 2016: Journal of Physical Chemistry. B
https://www.readbyqxmd.com/read/27737926/engineering-a-targeted-delivery-platform-using-centyrins
#9
Shalom D Goldberg, Rosa M F Cardoso, Tricia Lin, Tracy Spinka-Doms, Donna Klein, Steven A Jacobs, Vadim Dudkin, Gary Gilliland, Karyn T O'Neil
Targeted delivery of therapeutic payloads to specific tissues and cell types is an important component of modern pharmaceutical development. Antibodies or other scaffold proteins can provide the cellular address for delivering a covalently linked therapeutic via specific binding to cell-surface receptors. Optimization of the conjugation site on the targeting protein, linker chemistry and intracellular trafficking pathways can all influence the efficiency of delivery and potency of the drug candidate. In this study, we describe a comprehensive engineering experiment for an EGFR binding Centyrin, a highly stable fibronectin type III (FN3) domain, wherein all possible single-cysteine replacements were evaluated for expression, purification, conjugation efficiency, retention of target binding, biophysical properties and delivery of a cytotoxic small molecule payload...
December 2016: Protein Engineering, Design & Selection: PEDS
https://www.readbyqxmd.com/read/27729195/pyrazolo-1-5a-pyrimidines-as-a-new-class-of-fuse-binding-protein-1-fubp1-inhibitors
#10
Stefanie Hauck, Kerstin Hiesinger, Sabrina Khageh Hosseini, Janosch Achenbach, Ricardo M Biondi, Ewgenij Proschak, Martin Zörnig, Dalibor Odadzic
The transcriptional regulator FUSE binding protein 1 (FUBP1) is aberrantly upregulated in various malignancies, fulfilling its oncogenic role by the deregulation of critical genes involved in cell cycle control and apoptosis regulation. Thus, the pharmaceutical inhibition of this protein would represent an encouraging novel targeted chemotherapy. Here, we demonstrate the identification and initial optimization of a pyrazolo[1,5a]pyrimidine-based FUBP1 inhibitor derived from medium throughput screening, which interferes with the binding of FUBP1 to its single stranded target DNA FUSE...
September 14, 2016: Bioorganic & Medicinal Chemistry
https://www.readbyqxmd.com/read/27725965/tension-dependent-free-energies-of-nucleosome-unwrapping
#11
Joshua Lequieu, Andrés Córdoba, David C Schwartz, Juan J de Pablo
Nucleosomes form the basic unit of compaction within eukaryotic genomes, and their locations represent an important, yet poorly understood, mechanism of genetic regulation. Quantifying the strength of interactions within the nucleosome is a central problem in biophysics and is critical to understanding how nucleosome positions influence gene expression. By comparing to single-molecule experiments, we demonstrate that a coarse-grained molecular model of the nucleosome can reproduce key aspects of nucleosome unwrapping...
September 28, 2016: ACS Central Science
https://www.readbyqxmd.com/read/27721441/molecular-dynamics-simulations-and-docking-enable-to-explore-the-biophysical-factors-controlling-the-yields-of-engineered-nanobodies
#12
Miguel A Soler, Ario de Marco, Sara Fortuna
Nanobodies (VHHs) have proved to be valuable substitutes of conventional antibodies for molecular recognition. Their small size represents a precious advantage for rational mutagenesis based on modelling. Here we address the problem of predicting how Camelidae nanobody sequences can tolerate mutations by developing a simulation protocol based on all-atom molecular dynamics and whole-molecule docking. The method was tested on two sets of nanobodies characterized experimentally for their biophysical features...
October 10, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27714062/direct-real-time-detection-of-single-proteins-using-silicon-nanowire-based-electrical-circuits
#13
Jie Li, Gen He, Hiroshi Ueno, Chuancheng Jia, Hiroyuki Noji, Chuanmin Qi, Xuefeng Guo
We present an efficient strategy through surface functionalization to build a single silicon nanowire field-effect transistor-based biosensor that is capable of directly detecting protein adsorption/desorption at the single-event level. The step-wise signals in real-time detection of His-tag F1-ATPases demonstrate a promising electrical biosensing approach with single-molecule sensitivity, thus opening up new opportunities for studying single-molecule biophysics in broad biological systems.
September 15, 2016: Nanoscale
https://www.readbyqxmd.com/read/27677511/dynamic-and-active-proteins-biomolecular-motors-in-engineered-nanostructures
#14
Marisela Vélez
In Nature, proteins perform functions that go well beyond controlled self-assembly at the nano scale. They are the principal components of diverse "biological machines" that can self-assemble into dynamic aggregates that achieve the cold conversion of chemical energy into motion to realize complex functions involved in cell division, cellular transport and cell motility. Nowadays, we have identified many of the proteins involved in these "molecular machines" and know much about their biochemistry, structure and biophysical behavior...
2016: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/27676313/dna-based-nanopore-sensing
#15
Lei Liu, Hai-Chen Wu
Nanopore sensing is an attractive, label-free approach that can measure single molecules. Although initially proposed for rapid and low-cost DNA sequencing, nanopore sensors have been successfully employed in the detection of a wide variety of substrates. Early successes were mostly achieved based on two main strategies by 1) creating sensing elements inside the nanopore through protein mutation and chemical modification or 2) using molecular adapters to enhance analyte recognition. Over the past five years, DNA molecules started to be used as probes for sensing rather than substrates for sequencing...
September 27, 2016: Angewandte Chemie
https://www.readbyqxmd.com/read/27673570/tracking-transient-conformational-states-of-t4-lysozyme-at-room-temperature-combining-x-ray-crystallography-and-site-directed-spin-labeling
#16
Philipp Consentius, Ulrich Gohlke, Bernhard Loll, Claudia Alings, Robert Müller, Udo Heinemann, Martin Kaupp, Markus Wahl, Thomas Risse
Proteins are dynamic molecules that can transiently adopt different conformational states. As the function of the system often depends critically on its conformational state a rigorous understanding of the correlation between structure, energetics and dynamics of the different accessible states is crucial. The biophysical characterization of such processes is, however, challenging as the excited states are often only marginally populated. We show that a combination of X-ray crystallography performed at 100 K as well as at room temperature and EPR spectroscopy on a spin-labeled single crystal allows to correlate the structures of the ground state and a thermally excited state with their thermodynamics using the variant 118R1 of T4 lysozyme as an example...
October 5, 2016: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/27672145/single-molecule-imaging-of-electroporated-dye-labelled-chey-in-live-escherichia-coli
#17
Diana Di Paolo, Oshri Afanzar, Judith P Armitage, Richard M Berry
For the past two decades, the use of genetically fused fluorescent proteins (FPs) has greatly contributed to the study of chemotactic signalling in Escherichia coli including the activation of the response regulator protein CheY and its interaction with the flagellar motor. However, this approach suffers from a number of limitations, both biological and biophysical: for example, not all fusions are fully functional when fused to a bulky FP, which can have a similar molecular weight to its fused counterpart; they may interfere with the native interactions of the protein and the chromophores of FPs have low brightness and photostability and fast photobleaching rates...
November 5, 2016: Philosophical Transactions of the Royal Society of London. Series B, Biological Sciences
https://www.readbyqxmd.com/read/27671749/arachidonic-acid-mediates-the-formation-of-abundant-alpha-helical-multimers-of-alpha-synuclein
#18
Marija Iljina, Laura Tosatto, Minee L Choi, Jason C Sang, Yu Ye, Craig D Hughes, Clare E Bryant, Sonia Gandhi, David Klenerman
The protein alpha-synuclein (αS) self-assembles into toxic beta-sheet aggregates in Parkinson's disease, while it is proposed that αS forms soluble alpha-helical multimers in healthy neurons. Here, we have made αS multimers in vitro using arachidonic acid (ARA), one of the most abundant fatty acids in the brain, and characterized them by a combination of bulk experiments and single-molecule Fӧrster resonance energy transfer (sm-FRET) measurements. The data suggest that ARA-induced oligomers are alpha-helical, resistant to fibril formation, more prone to disaggregation, enzymatic digestion and degradation by the 26S proteasome, and lead to lower neuronal damage and reduced activation of microglia compared to the oligomers formed in the absence of ARA...
September 27, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27659445/quantum-entanglement-facts-and-fiction-how-wrong-was-einstein-after-all
#19
Bengt Nordén
Einstein was wrong with his 1927 Solvay Conference claim that quantum mechanics is incomplete and incapable of describing diffraction of single particles. However, the Einstein-Podolsky-Rosen paradox of entangled pairs of particles remains lurking with its 'spooky action at a distance'. In molecules quantum entanglement can be viewed as basis of both chemical bonding and excitonic states. The latter are important in many biophysical contexts and involve coupling between subsystems in which virtual excitations lead to eigenstates of the total Hamiltonian, but not for the separate subsystems...
January 2016: Quarterly Reviews of Biophysics
https://www.readbyqxmd.com/read/27658939/bridging-the-gap-between-in-vitro-and-in-vivo-rna-folding
#20
Kathleen A Leamy, Sarah M Assmann, David H Mathews, Philip C Bevilacqua
Deciphering the folding pathways and predicting the structures of complex three-dimensional biomolecules is central to elucidating biological function. RNA is single-stranded, which gives it the freedom to fold into complex secondary and tertiary structures. These structures endow RNA with the ability to perform complex chemistries and functions ranging from enzymatic activity to gene regulation. Given that RNA is involved in many essential cellular processes, it is critical to understand how it folds and functions in vivo...
January 2016: Quarterly Reviews of Biophysics
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