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https://www.readbyqxmd.com/read/28812882/aberrantly-large-single-channel-conductance-of-polyhistidine-arm-containing-protein-nanopores
#1
Avinash Kumar Thakur, Motahareh Ghahari Larimi, Kristin Gooden, Liviu Movileanu
There have been only a few studies reporting on the impact of polyhistidine affinity tags on the structure, function, and dynamics of proteins. Because of their relatively short size, they are often assumed to have little or no effect on the conformation and activity of a protein. Here, using membrane protein design and single-molecule electrophysiology, we determined that the presence of a hexahistidine arm at the N-terminus of a truncated FhuA-based protein nanopore, leaving the C-terminus untagged, produces an unusual increase in the unitary conductance up to ~8 nS in 1 M KCl...
August 16, 2017: Biochemistry
https://www.readbyqxmd.com/read/28812356/single-molecule-nanopore-spectrometry-for-peptide-detection
#2
Amy E Chavis, Kyle T Brady, Grace A Hatmaker, Christopher E Angevine, Nuwan Kothalawala, Amala Dass, Joseph W F Robertson, Joseph E Reiner
Sensing and characterization of water-soluble peptides is of critical importance in a wide variety of bioapplications. Single molecule nanopore spectrometry (SMNS) is based on the idea that one can use biological protein nanopores to resolve different sized molecules down to limits set by the blockade duration and noise. Previous work has shown that this enables discrimination between polyethylene glycol (PEG) molecules that differ by a single monomer unit. This paper describes efforts to extend SMNS to a variety of biologically relevant, water-soluble peptides...
August 16, 2017: ACS Sensors
https://www.readbyqxmd.com/read/28812314/self-assembled-chiral-nanostructures-of-amphiphilic-peptide-from-single-molecule-to-aggregate
#3
Ting Zhou, Zhiqing Zhang, Xuemei Zhang, Chen Wang, Guiying Xu, Yanlian Yang
We report interesting hierarchical self-assembled architectures from a designed amphiphilic peptide. The bisignate cotton effects in circular dichroism spectra show typical peptide aggregation-induced. The observation of peptide assembly structures from initial particles and fibrils to ribbon structures is supported by microscopy (atomic force microscopy and transmission electron microscopy). The visualization of individual peptide at the single molecular level offered insights of the intermolecular interactions responsible for the formation of aggregates, which is investigated by scanning tunneling microscopy...
August 15, 2017: Journal of Peptide Science: An Official Publication of the European Peptide Society
https://www.readbyqxmd.com/read/28811753/characterization-of-the-rosellinia-necatrix-transcriptome-and-genes-related-to-pathogenesis-by-single-molecule-mrna-sequencing
#4
Hyeongmin Kim, Seung Jae Lee, Ick-Hyun Jo, Jinsu Lee, Wonsil Bae, Hyemin Kim, Kyungho Won, Tae Kyung Hyun, Hojin Ryu
White root rot disease, caused by the pathogen Rosellinia necatrix, is one of the world's most devastating plant fungal diseases and affects several commercially important species of fruit trees and crops. Recent global outbreaks of R. necatrix and advances in molecular techniques have both increased interest in this pathogen. However, the lack of information regarding the genomic structure and transcriptome of R. necatrix has been a barrier to the progress of functional genomic research and the control of this harmful pathogen...
August 2017: Plant Pathology Journal
https://www.readbyqxmd.com/read/28809097/piconewton-scale-analysis-of-ras-braf-signal-transduction-with-single-molecule-force-spectroscopy
#5
Chae-Seok Lim, Cheng Wen, Yanghui Sheng, Guangfu Wang, Zhuan Zhou, Shiqiang Wang, Huaye Zhang, Anpei Ye, J Julius Zhu
Intermolecular interactions dominate the behavior of signal transduction in various physiological and pathological cell processes, yet assessing these interactions remains a challenging task. Here, this study reports a single-molecule force spectroscopic method that enables functional delineation of two interaction sites (≈35 pN and ≈90 pN) between signaling effectors Ras and BRaf in the canonical mitogen-activated protein kinase (MAPK) pathway. This analysis reveals mutations on BRaf at Q257 and A246, two sites frequently linked to cardio-faciocutaneous syndrome, result in ≈10-30 pN alterations in RasBRaf intermolecular binding force...
August 15, 2017: Small
https://www.readbyqxmd.com/read/28808695/resolving-multicopy-duplications-de-novo-using-polyploid-phasing
#6
Mark J Chaisson, Sudipto Mukherjee, Sreeram Kannan, Evan E Eichler
While the rise of single-molecule sequencing systems has enabled an unprecedented rise in the ability to assemble complex regions of the genome, long segmental duplications in the genome still remain a challenging frontier in assembly. Segmental duplications are at the same time both gene rich and prone to large structural rearrangements, making the resolution of their sequences important in medical and evolutionary studies. Duplicated sequences that are collapsed in mammalian de novo assemblies are rarely identical; after a sequence is duplicated, it begins to acquire paralog specific variants...
May 2017: Research in Computational Molecular Biology: ... Annual International Conference, RECOMB ...: Proceedings
https://www.readbyqxmd.com/read/28808686/a-conformational-checkpoint-between-dna-binding-and-cleavage-by-crispr-cas9
#7
Yavuz S Dagdas, Janice S Chen, Samuel H Sternberg, Jennifer A Doudna, Ahmet Yildiz
The Cas9 endonuclease is widely used for genome engineering applications by programming its single-guide RNA, and ongoing work is aimed at improving the accuracy and efficiency of DNA targeting. DNA cleavage of Cas9 is controlled by the conformational state of the HNH nuclease domain, but the mechanism that governs HNH activation at on-target DNA while reducing cleavage activity at off-target sites remains poorly understood. Using single-molecule Förster resonance energy transfer, we identified an intermediate state of Streptococcus pyogenes Cas9, representing a conformational checkpoint between DNA binding and cleavage...
August 2017: Science Advances
https://www.readbyqxmd.com/read/28808323/stress-hormone-rapidly-tunes-synaptic-nmda-receptor-through-membrane-dynamics-and-mineralocorticoid-signalling
#8
Lenka Mikasova, Hui Xiong, Amber Kerkhofs, Delphine Bouchet, Harm J Krugers, Laurent Groc
Stress hormones, such as corticosteroids, modulate the transmission of hippocampal glutamatergic synapses and NMDA receptor (NMDAR)-dependent synaptic plasticity, favouring salient behavioural responses to the environment. The corticosterone-induced synaptic adaptations partly rely on changes in NMDAR signalling, although the cellular pathway underlying this effect remains elusive. Here, we demonstrate, using single molecule imaging and electrophysiological approaches in hippocampal neurons, that corticosterone specifically controls GluN2B-NMDAR surface dynamics and synaptic content through mineralocorticoid signalling...
August 14, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28806726/replication-fork-slowing-and-stalling-are-distinct-checkpoint-independent-consequences-of-replicating-damaged-dna
#9
Divya Ramalingam Iyer, Nicholas Rhind
In response to DNA damage during S phase, cells slow DNA replication. This slowing is orchestrated by the intra-S checkpoint and involves inhibition of origin firing and reduction of replication fork speed. Slowing of replication allows for tolerance of DNA damage and suppresses genomic instability. Although the mechanisms of origin inhibition by the intra-S checkpoint are understood, major questions remain about how the checkpoint regulates replication forks: Does the checkpoint regulate the rate of fork progression? Does the checkpoint affect all forks, or only those encountering damage? Does the checkpoint facilitate the replication of polymerase-blocking lesions? To address these questions, we have analyzed the checkpoint in the fission yeast Schizosaccharomyces pombe using a single-molecule DNA combing assay, which allows us to unambiguously separate the contribution of origin and fork regulation towards replication slowing, and allows us to investigate the behavior of individual forks...
August 14, 2017: PLoS Genetics
https://www.readbyqxmd.com/read/28806521/determination-of-caspase-3-activity-and-its-inhibition-constant-by-combination-of-fluorescence-correlation-spectroscopy-with-a-microwell-chip
#10
Di Su, Xiaocai Hu, Chaoqing Dong, Jicun Ren
Caspase-3 is a key enzyme executing apoptosis during ontogenesis and homeostasis of multicellular organisms, and is a very important and potential drug target in treatment of apoptosis disturbance. So far, no commercial drugs for caspase-3 are available, and it is urgently necessitated to develop an effective method for caspase-3 activity assay and its inhibitor screening. In this paper, we propose a new method for determination of caspase-3 activity and its inhibition constant by combining single molecule fluorescence correlation spectroscopy (FCS) with a microwell chip...
August 14, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28806075/reversible-on-off-switching-of-a-single-molecule-magnet-via-a-crystal-to-crystal-chemical-transformation
#11
Dong-Qing Wu, Dong Shao, Xiao-Qin Wei, Fu-Xing Shen, Le Shi, David Kempe, Yuan-Zhu Zhang, Kim R Dunbar, Xin-Yi Wang
Dynamic molecular crystals are of great interest due to their potential applications. Herein we report the reversible on-off switching of single-molecule magnet (SMM) behavior in a [Mo(CN)7](4-) based molecular compound. Upon dehydration and rehydration, the trinuclear Mn2Mo molecule [Mn(L)(H2O)]2[Mo(CN)7]·2H2O (1) undergoes reversible crystal-to-crystal transformation to a hexanuclear Mn4Mo2 compound [Mn(L)(H2O)]2[Mn(L)]2[Mo(CN)7]2 (2). This structural transformation involves the breaking and reforming of coordination bonds which leads to significant changes in the color and magnetic properties...
August 14, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28805818/all-electric-control-of-donor-nuclear-spin-qubits-in-silicon
#12
Anthony J Sigillito, Alexei M Tyryshkin, Thomas Schenkel, Andrew A Houck, Stephen A Lyon
The electronic and nuclear spin degrees of freedom of donor impurities in silicon form ultra-coherent two-level systems that are potentially useful for applications in quantum information and are intrinsically compatible with industrial semiconductor processing. However, because of their smaller gyromagnetic ratios, nuclear spins are more difficult to manipulate than electron spins and are often considered too slow for quantum information processing. Moreover, although alternating current magnetic fields are the most natural choice to drive spin transitions and implement quantum gates, they are difficult to confine spatially to the level of a single donor, thus requiring alternative approaches...
August 14, 2017: Nature Nanotechnology
https://www.readbyqxmd.com/read/28805817/room-temperature-current-blockade-in-atomically-defined-single-cluster-junctions
#13
Giacomo Lovat, Bonnie Choi, Daniel W Paley, Michael L Steigerwald, Latha Venkataraman, Xavier Roy
Fabricating nanoscopic devices capable of manipulating and processing single units of charge is an essential step towards creating functional devices where quantum effects dominate transport characteristics. The archetypal single-electron transistor comprises a small conducting or semiconducting island separated from two metallic reservoirs by insulating barriers. By enabling the transfer of a well-defined number of charge carriers between the island and the reservoirs, such a device may enable discrete single-electron operations...
August 14, 2017: Nature Nanotechnology
https://www.readbyqxmd.com/read/28805060/substrate-induced-facilitated-dissociation-of-competitive-inhibitor-from-the-active-site-of-o-acetyl-serine-sulfhydrylase-reveal-a-competitive-allostery-mechanism
#14
Appu Kumar Singh, Mary Krishna Ekka, Abhishek Kaushik, Vaibhav Kumar Pandya, Ravi Pratap Singh, Srijita Banerjee, Monica Mittal, Vijay Singh, Sangaralingam Kumaran
By classical competitive-antagonism, substrate and competitive-inhibitor must bind mutually exclusively to the active site. The competitive inhibition of O-acetyl serine sulfhydrylase (OASS) by C-terminal of serine acetyltransferase (SAT) presents a paradox, because the C-terminal of SAT binds to the active site of OASS with 4-6 log fold (104 -106) more affinity than that of the substrate. Therefore, we employed multiple approaches to understand how substrate gains access to OASS active site under physiological conditions...
August 14, 2017: Biochemistry
https://www.readbyqxmd.com/read/28804000/the-mechanics-of-dna-loops-bridged-by-proteins-unveiled-by-single-molecule-experiments
#15
REVIEW
Catherine Tardin
Protein-induced DNA bridging and looping is a common mechanism for various and essential processes in bacterial chromosomes. This mechanism is preserved despite the very different bacterial conditions and their expected influence on the thermodynamic and kinetic characteristics of the bridge formation and stability. Over the last two decades, single-molecule techniques carried out on in vitro DNA systems have yielded valuable results which, in combination with theoretical works, have clarified the effects of different parameters of nucleoprotein complexes on the protein-induced DNA bridging and looping process...
August 10, 2017: Biochimie
https://www.readbyqxmd.com/read/28803781/myc-regulates-chromatin-decompaction-and-nuclear-architecture-during-b-cell-activation
#16
Kyong-Rim Kieffer-Kwon, Keisuke Nimura, Suhas S P Rao, Jianliang Xu, Seolkyoung Jung, Aleksandra Pekowska, Marei Dose, Evan Stevens, Ewy Mathe, Peng Dong, Su-Chen Huang, Maria Aurelia Ricci, Laura Baranello, Ying Zheng, Francesco Tomassoni Ardori, Wolfgang Resch, Diana Stavreva, Steevenson Nelson, Michael McAndrew, Adriel Casellas, Elizabeth Finn, Charles Gregory, Brian Glenn St Hilaire, Steven M Johnson, Wendy Dubois, Maria Pia Cosma, Eric Batchelor, David Levens, Robert D Phair, Tom Misteli, Lino Tessarollo, Gordon Hager, Melike Lakadamyali, Zhe Liu, Monique Floer, Hari Shroff, Erez Lieberman Aiden, Rafael Casellas
50 years ago, Vincent Allfrey and colleagues discovered that lymphocyte activation triggers massive acetylation of chromatin. However, the molecular mechanisms driving epigenetic accessibility are still unknown. We here show that stimulated lymphocytes decondense chromatin by three differentially regulated steps. First, chromatin is repositioned away from the nuclear periphery in response to global acetylation. Second, histone nanodomain clusters decompact into mononucleosome fibers through a mechanism that requires Myc and continual energy input...
July 28, 2017: Molecular Cell
https://www.readbyqxmd.com/read/28802172/interaction-of-phenazinium-based-photosensitizers-with-the-n-and-b-isoforms-of-human-serum-albumin-effect-of-methyl-substitution
#17
Swagata Sen, Riya Sett, Bijan K Paul, Nikhil Guchhait
The present work is focused on exploring the interaction of two phenazinium-based biological photosensitizers, phenosafranin (PSF) and safranin-O (SO), with human serum albumin (HSA), with particular emphasis on the physiologically significant NB conformational transition of the protein on the dye:HSA interaction. In addition, the presence of methyl substitution on the planar phenazinium ring in SO paves way for looking into the effect of simple chemical manipulation (that is, methyl substitution on the dye nucleus) on the dye:protein interaction behavior as a function of various (pH-induced) isoforms of HSA...
August 3, 2017: Journal of Photochemistry and Photobiology. B, Biology
https://www.readbyqxmd.com/read/28801929/validation-and-application-of-a-novel-integrated-genetic-screening-method-to-a-cohort-of-1-112-men-with-idiopathic-azoospermia-or-severe-oligozoospermia
#18
Manon S Oud, Liliana Ramos, Moira K O'Bryan, Robert I McLachlan, Özlem Okutman, Stephane Viville, Petra F de Vries, Dominique F C M Smeets, Dorien Lugtenberg, Jayne Y Hehir-Kwa, Christian Gilissen, Maartje van de Vorst, Lisenka E L M Vissers, Alexander Hoischen, Aukje M Meijerink, Kathrin Fleischer, Joris A Veltman, Michiel J Noordam
Microdeletions of the Y chromosome (YCMs), Klinefelter syndrome (47,XXY) and CFTR mutations are known genetic causes of severe male infertility, but the majority of cases remain idiopathic. Here we describe a novel method using single molecule Molecular Inversion Probes (smMIPs), to screen infertile men for mutations and copy number variations (CNVs) affecting known disease genes. We designed a set of 4,525 smMIPs targeting the coding regions of causal (n = 6) and candidate (n = 101) male infertility genes...
August 11, 2017: Human Mutation
https://www.readbyqxmd.com/read/28800752/a-type-i-combi-targeting-approach-for-the-design-of-molecules-with-enhanced-potency-against-brca1-2-mutant-and-o6-methylguanine-dna-methyltransferase-mgmt-expressing-tumour-cells
#19
Zhor Senhaji Mouhri, Elliot Goodfellow, Bertrand Jean-Claude
BACKGROUND: Mutations of the DNA repair proteins BRCA1/2 are synthetically lethal with the DNA repair enzyme poly(ADP-ribose) polymerase (PARP), which when inhibited, leads to cell death due to the absence of compensatory DNA repair mechanism. The potency of PARP inhibitors has now been clinically proven. However, disappointingly, acquired resistance mediated by the reactivation of wild type BRCA1/2 has been reported. In order to improve their efficacy, trials are ongoing to explore their combinations with temozolomide (TMZ)...
August 11, 2017: BMC Cancer
https://www.readbyqxmd.com/read/28800289/viral-master-circuits-from-discovery-to-new-therapy-targets
#20
Anand Pai, Leor S Weinberger
Current antivirals effectively target diverse viruses at various stages of their life cycles. Nevertheless, curative therapy has remained elusive for important pathogens, such as human immunodeficiency virus type 1 (HIV-1) and herpesviruses, in large part due to viral latency and the evolution of resistance to existing therapies. Here, we review the discovery of viral master circuits: virus-encoded autoregulatory gene networks that autonomously control viral expression programs (i.e., between active, latent, and abortive fates)...
August 11, 2017: Annual Review of Virology
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