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Protein footprinting

Benjamin B Minkoff, Joshua M Blatz, Faraz A Choudhury, Daniel Benjamin, J Leon Shohet, Michael R Sussman
Protein three-dimensional structure dynamically changes in solution depending on the presence of ligands and interacting proteins. Methods for detecting these changes in protein conformation include 'protein footprinting,' using mass spectrometry. We describe herein a new technique, PLIMB (Plasma Induced Modification of Biomolecules), that generates µs bursts of hydroxyl radicals from water, to measure changes in protein structure via altered solvent accessibility of amino acid side chains. PLIMB was first benchmarked with model compounds, and then applied to a biological problem, i...
October 11, 2017: Scientific Reports
Yoram Zarai, Michael Margaliot, Tamir Tuller
We study a deterministic mechanistic model for the flow of ribosomes along the mRNA molecule, called the ribosome flow model with extended objects (RFMEO). This model encapsulates many realistic features of translation including non-homogeneous transition rates along mRNA, the fact that every ribosome covers several codons, and the fact that ribosomes cannot overtake one another. The RFMEO is a mean-field approximation of an important model from statistical mechanics called the totally asymmetric simple exclusion process with extended objects (TASEPEO)...
October 2017: Journal of the Royal Society, Interface
Marina Cavaiuolo, Richard Kuras, Francis-André Wollman, Yves Choquet, Olivier Vallon
In Chlamydomonas reinhardtii, regulation of chloroplast gene expression is mainly post-transcriptional. It requires nucleus-encoded trans-acting protein factors for maturation/stabilization (M factors) or translation (T factors) of specific target mRNAs. We used long- and small-RNA sequencing to generate a detailed map of the transcriptome. Clusters of sRNAs marked the 5' end of all mature mRNAs. Their absence in M-factor mutants reflects the protection of transcript 5' end by the cognate factor. Enzymatic removal of 5'-triphosphates allowed identifying those cosRNA that mark a transcription start site...
August 2, 2017: Nucleic Acids Research
Michael G Kearse, Jeremy E Wilusz
Although it was long thought that eukaryotic translation almost always initiates at an AUG start codon, recent advancements in ribosome footprint mapping have revealed that non-AUG start codons are used at an astonishing frequency. These non-AUG initiation events are not simply errors but instead are used to generate or regulate proteins with key cellular functions; for example, during development or stress. Misregulation of non-AUG initiation events contributes to multiple human diseases, including cancer and neurodegeneration, and modulation of non-AUG usage may represent a novel therapeutic strategy...
September 1, 2017: Genes & Development
Elin Teppa, Diego Javier Zea, Cristina Marino-Buslje
Protein-protein interactions are essential to all aspects of life. Specific interactions result from evolutionary pressure at the interacting interfaces of partner proteins. However, evolutionary pressure is not homogeneous within the interface: for instance, each residue does not contribute equally to the binding energy of the complex. To understand functional differences between residues within the interface, we analyzed their properties in the core and rim regions. Here, we characterized protein interfaces with two evolutionary measures, conservation and coevolution, using a comprehensive dataset of 896 protein complexes...
October 4, 2017: Protein Science: a Publication of the Protein Society
Taichi Umeyama, Takashi Ito
Protein-DNA interactions provide the basis for chromatin structure and gene regulation. Comprehensive identification of protein-occupied sites is thus vital to an in-depth understanding of genome function. Dimethyl sulfate (DMS) is a chemical probe that has long been used to detect footprints of DNA-bound proteins in vitro and in vivo. Here, we describe a genomic footprinting method, dimethyl sulfate sequencing (DMS-seq), which exploits the cell-permeable nature of DMS to obviate the need for nuclear isolation...
October 3, 2017: Cell Reports
Zachary Berndsen, Charles Bowman, Haerin Jang, Andrew B Ward
Summary: The Electron Microscopy Hole Punch (EMHP) is a streamlined suite of tools for quick assessment, sorting, and hole masking of electron micrographs. With recent advances in single-particle electron cryo-microscopy (cryo-EM) data processing allowing for the rapid determination of protein structures using a smaller computational footprint, we saw the need for a fast and simple tool for data pre-processing that could run independent of existing high-performance computing (HPC) infrastructures...
August 7, 2017: Bioinformatics
Lucio Manzi, Andrew Barrow, Jonathan Hopper, Renata Kaminska, Colin Kleanthous, Carol Robinson, John Moses, Neil J Oldham
Mapping the interaction sites between membrane spanning proteins is a key challenge in structural biology. In this study a carbene footprinting approach is developed and applied to identify the interfacial sites of a trimeric, integral membrane protein, OmpF, solubilised in micelles. The diazirine-based footprinting probe is effectively sequestered by, and incorporated into, the micelles leading to efficient labelling of the membrane-spanning regions of the protein upon irradiation at 349 nm. Areas associated with protein-protein interactions between the trimer subunits remained unlabelled, thus revealing their location...
September 27, 2017: Angewandte Chemie
Georgia G Kournoutou, Panagiota C Giannopoulou, Eleni Sazakli, Michel Leotsinidis, Dimitrios L Kalpaxis
Numerous studies have shown the ability of trace metals to accumulate in marine organisms and cause oxidative stress that leads to perturbations in many important intracellular processes, including protein synthesis. This study is mainly focused on the exploration of structural changes, like base modifications, scissions, and conformational changes, caused in 18S and 5S ribosomal RNA (rRNA) isolated from the mussel Mytilus galloprovincialis exposed to 40μg/L Cu, 30μg/L Hg, or 100μg/L Cd, for 5 or 15days...
September 6, 2017: Aquatic Toxicology
Aleksandra Binek, Rodrigo Fernández-Jiménez, Inmaculada Jorge, Emilio Camafeita, Juan Antonio López, Navratan Bagwan, Carlos Galán-Arriola, Andres Pun, Jaume Agüero, Valentin Fuster, Borja Ibanez, Jesús Vázquez
Reperfusion alters post-myocardial infarction (MI) healing; however, very few systematic studies report the early molecular changes following ischemia/reperfusion (I/R). Alterations in the remote myocardium have also been neglected, disregarding its contribution to post-MI heart failure (HF) development. This study characterizes protein dynamics and contractile abnormalities in the ischemic and remote myocardium during one week after MI. Closed-chest 40 min I/R was performed in 20 pigs sacrificed at 120 min, 24 hours, 4days, and 7days after reperfusion (n = 5 per group)...
September 27, 2017: Scientific Reports
Rainer Kluger, Klaus R Huber, Philipp G Seely, Christian E Berger, Florian Frommlet
BACKGROUND: Several single-nucleotide polymorphisms (SNPs) in the TNC gene have recently been found to be associated with degenerative rotator cuff tears. HYPOTHESIS: Exonic SNPs in the TNC gene are related to the risk for a failure to heal after rotator cuff repair. STUDY DESIGN: Case-control study; Level of evidence, 3. METHODS: A total of 302 patients from the Vienna area and European Caucasian ancestry underwent mini-open rotator cuff repair for a full-thickness superior or posterosuperior tear and were assessed for the integrity of the repair 1 year postoperatively with a real-time 7...
September 1, 2017: American Journal of Sports Medicine
Alexey K Shaytan, Hua Xiao, Grigoriy A Armeev, Carl Wu, David Landsman, Anna R Panchenko
Nucleosomes are the most abundant protein-DNA complexes in eukaryotes that provide compaction of genomic DNA and are implicated in regulation of transcription, DNA replication and repair. The details of DNA positioning on the nucleosome and the DNA conformation can provide key regulatory signals. Hydroxyl-radical footprinting (HRF) of protein-DNA complexes is a chemical technique that probes nucleosome organization in solution with a high precision unattainable by other methods. In this work we propose an integrative modeling method for constructing high-resolution atomistic models of nucleosomes based on HRF experiments...
September 19, 2017: Nucleic Acids Research
Parwinder Kaur, Rudi Appels, Philipp E Bayer, Gabriel Keeble-Gagnere, Jiankang Wang, Hideki Hirakawa, Kenta Shirasawa, Philip Vercoe, Katia Stefanova, Zoey Durmic, Phillip Nichols, Clinton Revell, Sachiko N Isobe, David Edwards, William Erskine
Mitigating methane production by ruminants is a significant challenge to global livestock production. This research offers a new paradigm to reduce methane emissions from ruminants by breeding climate-clever clovers. We demonstrate wide genetic diversity for the trait methanogenic potential in Australia's key pasture legume, subterranean clover (Trifolium subterraneum L.). In a bi-parental population the broadsense heritability in methanogenic potential was moderate (H(2) = 0.4) and allelic variation in a region of Chr 8 accounted for 7...
2017: Frontiers in Plant Science
Helen Yakhnin, Robert Aichele, Sarah E Ades, Tony Romeo, Paul Babitzke
CsrA of Escherichia coli is an RNA-binding protein that globally regulates a wide variety of cellular processes and behaviors including carbon metabolism, motility, biofilm formation, and the stringent response. CsrB and CsrC are sRNAs that sequester CsrA, thereby preventing CsrA-mRNA interaction. RpoE (σ(E)) is the extracytoplasmic stress response sigma factor of E. coli Previous RNA-seq studies identified rpoE mRNA as a CsrA target. Here we explored the regulation of rpoE by CsrA and found that CsrA represses rpoE translation...
September 18, 2017: Journal of Bacteriology
Pascale Jolivet, Laure Aymé, Alexandre Giuliani, Frank Wien, Thierry Chardot, Yann Gohon
Lipid droplets are the major stock of lipids in oleaginous plant seeds. Despite their economic importance for oil production and biotechnological issues (biofuels, lubricants and plasticizers), numerous questions about their formation, structure and regulation are still unresolved. To determine water accessible domains of protein coating at lipid droplets surface, a structural proteomic approach has been performed. This technique relies on the millisecond timescale production of hydroxyl radicals by the radiolysis of water using Synchrotron X-ray white beam...
September 14, 2017: Journal of Proteomics
Brian K Dalley, Lisa Baird, Michael T Howard
Deep sequencing of ribosome protected mRNA footprints, also called ribosome profiling or Ribo-Seq, is a relatively new methodology well suited to address questions regarding the mechanisms and efficiency of protein expression. Specifically, the ability of this technique to quantify ribosome abundance with codon resolution enables experiments aimed at studying many aspects of translation, including gene-specific translational efficiency, translation of regulatory upstream short open reading frames, sites of ribosome pausing, and most importantly for selenoproteins, the efficiency by which UGA codons are redefined to encode selenocysteine...
2018: Methods in Molecular Biology
Azusa Oita, Ichiro Nagano, Hiroyuki Matsuda
Dietary choices largely affect human-induced reactive nitrogen accumulation in the environment and resultant environmental problems. A nitrogen footprint (NF) is an indicator of how an individual's consumption patterns impact nitrogen pollution. Here, we examined the impact of changes in the Japanese diet from 1961 to 2011 and the effect of alternative diets (the recommended protein diet, a pescetarian diet, a low-NF food diet, and a balanced Japanese diet) on the food NF. The annual per capita Japanese food NF has increased by 55% as a result of dietary changes since 1961...
September 14, 2017: Ambio
Jarno Drost, Ruben van Boxtel, Francis Blokzijl, Tomohiro Mizutani, Nobuo Sasaki, Valentina Sasselli, Joep de Ligt, Sam Behjati, Judith E Grolleman, Tom van Wezel, Serena Nik-Zainal, Roland P Kuiper, Edwin Cuppen, Hans Clevers
Mutational processes underlie cancer initiation and progression. Signatures of these processes in cancer genomes may explain cancer etiology and could hold diagnostic and prognostic value. We developed a strategy that can be used to explore the origin of cancer-associated mutational signatures. We used CRISPR-Cas9 technology to delete key DNA repair genes in human colon organoids, followed by delayed subcloning and whole-genome sequencing. We found that mutation accumulation in organoids deficient in the mismatch repair gene MLH1 is driven by replication errors and accurately models the mutation profiles observed in mismatch repair-deficient colorectal cancers...
October 13, 2017: Science
Christopher Lockhart, Dmitri K Klimov
Using isobaric-isothermal all-atom replica-exchange molecular dynamics (REMD) simulations, we investigated the equilibrium binding of Aβ10-40 monomers to the zwitterionic dimyristoylphosphatidylcholine (DMPC) bilayer containing cholesterol. Our previous REMD simulations, which studied binding of the same peptide to the cholesterol-free DMPC bilayer, served as a control, against which we measured the impact of cholesterol. Our findings are as follows. First, addition of cholesterol to the DMPC bilayer partially expels the Aβ peptide from the hydrophobic core and promotes its binding to bilayer polar headgroups...
October 2, 2017: Journal of Chemical Information and Modeling
Ron Schwessinger, Maria C Suciu, Simon J McGowan, Jelena Telenius, Stephen Taylor, Doug R Higgs, Jim R Hughes
In the era of genome-wide association studies (GWAS) and personalized medicine, predicting the impact of single nucleotide polymorphisms (SNPs) in regulatory elements is an important goal. Current approaches to determine the potential of regulatory SNPs depend on inadequate knowledge of cell-specific DNA binding motifs. Here, we present Sasquatch, a new computational approach that uses DNase footprint data to estimate and visualize the effects of noncoding variants on transcription factor binding. Sasquatch performs a comprehensive k-mer-based analysis of DNase footprints to determine any k-mer's potential for protein binding in a specific cell type and how this may be changed by sequence variants...
October 2017: Genome Research
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