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Single molecule biophysics

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https://www.readbyqxmd.com/read/28315749/integrated-structural-biology-to-unravel-molecular-mechanisms-of-protein-rna-recognition
#1
REVIEW
Andreas Schlundt, Jan-Niklas Tants, Michael Sattler
Recent advances in RNA sequencing technologies have greatly expanded our knowledge of the RNA landscape in cells, often with spatiotemporal resolution. These techniques identified many new (often non-coding) RNA molecules. Large-scale studies have also discovered novel RNA binding proteins (RBPs), which exhibit single or multiple RNA binding domains (RBDs) for recognition of specific sequence or structured motifs in RNA. Starting from these large-scale approaches it is crucial to unravel the molecular principles of protein-RNA recognition in ribonucleoprotein complexes (RNPs) to understand the underlying mechanisms of gene regulation...
March 15, 2017: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/28289210/phosphorylation-induced-conformational-dynamics-in-an-intrinsically-disordered-protein-and-potential-role-in-phenotypic-heterogeneity
#2
Prakash Kulkarni, Mohit Kumar Jolly, Dongya Jia, Steven M Mooney, Ajay Bhargava, Luciane T Kagohara, Yihong Chen, Pengyu Hao, Yanan He, Robert W Veltri, Alexander Grishaev, Keith Weninger, Herbert Levine, John Orban
Intrinsically disordered proteins (IDPs) that lack a unique 3D structure and comprise a large fraction of the human proteome play important roles in numerous cellular functions. Prostate-Associated Gene 4 (PAGE4) is an IDP that acts as a potentiator of the Activator Protein-1 (AP-1) transcription factor. Homeodomain-Interacting Protein Kinase 1 (HIPK1) phosphorylates PAGE4 at S9 and T51, but only T51 is critical for its activity. Here, we identify a second kinase, CDC-Like Kinase 2 (CLK2), which acts on PAGE4 and hyperphosphorylates it at multiple S/T residues, including S9 and T51...
March 13, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28287314/photonic-and-plasmonic-nanotweezing-of-nano-and-microscale-particles
#3
Donato Conteduca, Francesco Dell'Olio, Thomas F Krauss, Caterina Ciminelli
The ability to manipulate and sense biological molecules is important in many life science domains, such as single-molecule biophysics, the development of new drugs and cancer detection. Although the manipulation of biological matter at the nanoscale continues to be a challenge, several types of nanotweezers based on different technologies have recently been demonstrated to address this challenge. In particular, photonic and plasmonic nanotweezers are attracting a strong research effort especially because they are efficient and stable, they offer fast response time, and avoid any direct physical contact with the target object to be trapped, thus preventing its disruption or damage...
March 2017: Applied Spectroscopy
https://www.readbyqxmd.com/read/28256724/a-biophysical-view-on-von-willebrand-factor-activation
#4
REVIEW
Achim Löf, Jochen P Müller, Maria A Brehm
The process of hemostatic plug formation at sites of vascular injury crucially relies on the large multimeric plasma glycoprotein von Willebrand factor (VWF) and its ability to recruit platelets to the damaged vessel wall via interaction of its A1 domain with platelet GPIbα. Under normal blood flow conditions, VWF multimers exhibit a very low binding affinity for platelets. Only when subjected to increased hydrodynamic forces, which primarily occur in connection with vascular injury, VWF can efficiently bind to platelets...
March 3, 2017: Journal of Cellular Physiology
https://www.readbyqxmd.com/read/28255705/f%C3%A3-rster-resonance-energy-transfer-to-study-tcr-pmhc-interactions-in-the-immunological-synapse
#5
Gerhard J Schütz, Johannes B Huppa
T-cell antigen recognition is remarkably efficient: when scanning the surface of antigen-presenting cells (APCs), T-cells can detect the presence of just a few single antigenic peptide/MHCs (pMHCs), which are often vastly outnumbered by structurally similar non-stimulatory endogenous pMHCs (Irvine et al., Nature 419(6909):845-849, 2002; Purbhoo et al., Nat Immunol 5(5):524-530, 2004; Huang et al., Immunity 39(5):846-857, 2013). How T-cells achieve this is still enigmatic, in particular in view of the rather moderate affinity that TCRs typically exert for antigenic pMHCs, at least when measured in vitro (Davis et al...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28247968/breaking-up-and-making-up-the-secret-life-of-the-vacuolar-h-atpase
#6
REVIEW
Rebecca A Oot, Sergio Couoh-Cardel, Stuti Sharma, Nicholas J Stam, Stephan Wilkens
The vacuolar ATPase (V-ATPase; V1 Vo -ATPase) is a large multisubunit proton pump found in the endomembrane system of all eukaryotic cells where it acidifies the lumen of subcellular organelles including lysosomes, endosomes, the Golgi apparatus and clathrin coated vesicles. V-ATPase function is essential for pH and ion homeostasis, protein trafficking, endocytosis, mechanistic target of rapamycin (mTOR) and Notch signaling, as well as hormone secretion and neurotransmitter release. V-ATPase can also be found in the plasma membrane of polarized animal cells where its proton pumping function is involved in bone remodeling, urine acidification and sperm maturation...
March 1, 2017: Protein Science: a Publication of the Protein Society
https://www.readbyqxmd.com/read/28223017/novel-bcl2-inhibitor-disarib-induces-apoptosis-by-disruption-of-bcl2-bak-interaction
#7
Supriya V Vartak, Divyaanka Iyer, T R Santhoshkumar, Sheetal Sharma, Archita Mishra, Gunaseelan Goldsmith, Mrinal Srivastava, Shikha Srivastava, Subhas S Karki, Avadhesha Surolia, Bibha Choudhary, Sathees C Raghavan
Apoptosis is a highly regulated pathway of programmed cell death relying on the fine balance between pro and antiapoptotic binding partners. Overexpression of the antiapoptotic protein BCL2 in several cancers makes it an ideal target for chemotherapy, with minimum side effects. In one of our previous studies, we designed, synthesized and characterized Disarib, a BCL2-specific small molecule inhibitor. Interestingly, Disarib showed a novel mode of BCL2 inhibition, by predominantly binding to its BH1 domain, as compared to the BH3-specific action of other known BCL2 inhibitors...
February 20, 2017: Biochemical Pharmacology
https://www.readbyqxmd.com/read/28222010/optical-tweezers-studies-of-transcription-by-eukaryotic-rna-polymerases
#8
Ana Lisica, Stephan W Grill
Transcription is the first step in the expression of genetic information and it is carried out by large macromolecular enzymes called RNA polymerases. Transcription has been studied for many years and with a myriad of experimental techniques, ranging from bulk studies to high-resolution transcript sequencing. In this review, we emphasise the advantages of using single-molecule techniques, particularly optical tweezers, to study transcription dynamics. We give an overview of the latest results in the single-molecule transcription field, focusing on transcription by eukaryotic RNA polymerases...
March 1, 2017: Biomolecular Concepts
https://www.readbyqxmd.com/read/28210970/a-suspended-carbon-fiber-culture-to-model-myelination-by-human-schwann-cells
#9
Antonio Merolli, Yong Mao, Joachim Kohn
Understanding of myelination/remyelination process is essential to guide tissue engineering for nerve regeneration. In vitro models currently used are limited to cell population studies and cannot easily identify individual cell contribution to the process. We established a novel model to study the contribution of human Schwann cells to the myelination process. The model avoids the presence of neurons in culture; Schwann cells respond solely to the biophysical properties of an artificial axon. The model uses a single carbon fiber suspended in culture media far from the floor of the well...
April 2017: Journal of Materials Science. Materials in Medicine
https://www.readbyqxmd.com/read/28198806/arrays-of-individual-dna-molecules-on-nanopatterned-substrates
#10
Roland Hager, Alma Halilovic, Jonathan R Burns, Friedrich Schäffler, Stefan Howorka
Arrays of individual molecules can combine the advantages of microarrays and single-molecule studies. They miniaturize assays to reduce sample and reagent consumption and increase throughput, and additionally uncover static and dynamic heterogeneity usually masked in molecular ensembles. However, realizing single-DNA arrays must tackle the challenge of capturing structurally highly dynamic strands onto defined substrate positions. Here, we create single-molecule arrays by electrostatically adhering single-stranded DNA of gene-like length onto positively charged carbon nanoislands...
February 13, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28137359/pharmacological-targeting-of-the-transcription-factor-sox18-delays-breast-cancer-in-mice
#11
Jeroen Overman, Frank Fontaine, Mehdi Moustaqil, Deepak Mittal, Emma Sierecki, Natalia Sacilotto, Johannes Zuegg, Avril Ab Robertson, Kelly Holmes, Angela A Salim, Sreeman Mamidyala, Mark S Butler, Ashley S Robinson, Emmanuelle Lesieur, Wayne Johnston, Kirill Alexandrov, Brian L Black, Benjamin M Hogan, Sarah De Val, Robert J Capon, Jason S Carroll, Timothy L Bailey, Peter Koopman, Ralf Jauch, Mark J Smyth, Matthew A Cooper, Yann Gambin, Mathias Francois
Pharmacological targeting of transcription factors holds great promise for the development of new therapeutics, but strategies based on blockade of DNA binding, nuclear shuttling, or individual protein partner recruitment have yielded limited success to date. Transcription factors typically engage in complex interaction networks, likely masking the effects of specifically inhibiting single protein-protein interactions. Here, we used a combination of genomic, proteomic and biophysical methods to discover a suite of protein-protein interactions involving the SOX18 transcription factor, a known regulator of vascular development and disease...
January 31, 2017: ELife
https://www.readbyqxmd.com/read/28117936/protein-templated-fragment-ligations-from-molecular-recognition-to-drug-discovery
#12
Mike Jaegle, Ee Lin Wong, Carolin Tauber, Eric Nawrotzky, Christoph Arkona, Jörg Rademann
The understanding and manipulation of molecular recognition events is the key to modern approaches in drug discovery. Protein-templated fragment ligation is a novel concept to support drug discovery and can help to improve the efficacy of already existing protein ligands. Protein-templated fragment ligations are chemical reactions between small molecules ("fragments") that utilize a protein´s surface as a template to combine and to form a protein ligand with increased binding affinity. The approach exploits the molecular recognition of reactive small molecule fragments by proteins both for ligand assembly and for the identification of bioactive fragment combinations...
January 24, 2017: Angewandte Chemie
https://www.readbyqxmd.com/read/28092942/advances-in-structural-and-single-molecule-methods-for-investigating-dna-lesion-bypass-and-repair-polymerases
#13
Austin T Raper, Andrew J Reed, Varun V Gadkari, Zucai Suo
Innovative advances in X-ray crystallography and single-molecule biophysics have yielded unprecedented insight into the mechanisms of DNA lesion bypass and damage repair. Time-dependent X-ray crystallography has been successfully applied to view the bypass of 8-oxo-7,8-dihydro-2'-deoxyguanine (8-oxoG), a major oxidative DNA lesion, and the incorporation of the triphosphate form, 8-oxo-dGTP, catalyzed by human DNA polymerase β. Significant findings of these studies are highlighted here, and their contributions to the current mechanistic understanding of mutagenic translesion DNA synthesis (TLS) and base excision repair are discussed...
January 17, 2017: Chemical Research in Toxicology
https://www.readbyqxmd.com/read/28087691/imaging-proteins-at-the-single-molecule-level
#14
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...
February 14, 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
#15
REVIEW
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
#16
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 24, 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
#17
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
#18
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
#19
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
#20
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
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