scholarly journals XPRESSyourself: Enhancing, standardizing, and automating ribosome profiling computational analyses yields improved insight into data

2020 ◽  
Vol 16 (1) ◽  
pp. e1007625 ◽  
Author(s):  
Jordan A. Berg ◽  
Jonathan R. Belyeu ◽  
Jeffrey T. Morgan ◽  
Yeyun Ouyang ◽  
Alex J. Bott ◽  
...  
Keyword(s):  
Ribosome Profiling ◽  
Computational Analyses ◽  
Insight Into

scholarly journals XPRESSyourself: Enhancing, Standardizing, and Automating Ribosome Profiling Computational Analyses Yields Improved Insight into Data

2019 ◽  
Author(s):  
Jordan A. Berg ◽  
Jonathan R. Belyeu ◽  
Jeffrey T. Morgan ◽  
Yeyun Ouyang ◽  
Alex J. Bott ◽  
...  
Keyword(s):  
Best Practices ◽  
Protein Translation ◽  
Ribosome Profiling ◽  
Translation Regulation ◽  
Computational Tools ◽  
Current State ◽  
Reference Implementation ◽  
Computational Analyses ◽  
Profiling Analysis ◽  
Insight Into

AbstractRibosome profiling, an application of nucleic acid sequencing for monitoring ribosome activity, has revolutionized our understanding of protein translation dynamics. This technique has been available for a decade, yet the current state and standardization of publicly available computational tools for these data is bleak. We introduce XPRESSyourself, an analytical toolkit that eliminates barriers and bottlenecks associated with this specialized data type by filling gaps in the computational toolset for both experts and non-experts of ribosome profiling. XPRESSyourself automates and standardizes analysis procedures, decreasing time-to-discovery and increasing reproducibility. This toolkit acts as a reference implementation of current best practices in ribosome profiling analysis. We demonstrate this toolkit’s performance on publicly available ribosome profiling data by rapidly identifying hypothetical mechanisms related to neurodegenerative phenotypes and neuroprotective mechanisms of the small-molecule ISRIB during acute cellular stress. XPRESSyourself brings robust, rapid analysis of ribosome-profiling data to a broad and ever-expanding audience and will lead to more reproducible and accessible measurements of translation regulation. XPRESSyourself software is perpetually open-source under the GPL-3.0 license and is hosted at https://github.com/XPRESSyourself, where users can access additional documentation and report software issues.

scholarly journals Insight into Inhibitor Binding in the Eukaryotic Proteasome: Computations of the 20S CP

2018 ◽  
Vol 19 (12) ◽  
pp. 3858
Author(s):  
Milan Hodošček ◽  
Nadia Elghobashi-Meinhardt
Keyword(s):  
Electrostatic Interactions ◽  
Active Sites ◽  
Sampling Period ◽  
Md Simulations ◽  
Structural Features ◽  
Relaxation Dynamics ◽  
Inhibitor Binding ◽  
Computational Analyses ◽  
Insight Into ◽  
Proteolytic Chamber

A combination of molecular dynamics (MD) simulations and computational analyses uncovers structural features that may influence substrate passage and exposure to the active sites within the proteolytic chamber of the 20S proteasome core particle (CP). MD simulations of the CP reveal relaxation dynamics in which the CP slowly contracts over the 54 ns sampling period. MD simulations of the SyringolinA (SylA) inhibitor within the proteolytic B 1 ring chamber of the CP indicate that favorable van der Waals and electrostatic interactions account for the predominant association of the inhibitor with the walls of the proteolytic chamber. The time scale required for the inhibitor to travel from the center of the proteolytic chamber to the chamber wall is on the order of 4 ns, accompanied by an average energetic stabilization of approximately −20 kcal/mol.

scholarly journals Beetle luciferases with naturally red- and blue-shifted emission

2018 ◽  
Vol 1 (4) ◽  
pp. e201800072 ◽  
Author(s):  
César Carrasco-López ◽  
Juliana C Ferreira ◽  
Nathan M Lui ◽  
Stefan Schramm ◽  
Romain Berraud-Pache ◽  
...  
Keyword(s):  
Active Site ◽  
Computational Studies ◽  
Reaction Sequence ◽  
Yellow Light ◽  
Conformational Landscape ◽  
The Common ◽  
Molecular Origin ◽  
Site Mutagenesis ◽  
Computational Analyses ◽  
Insight Into

The different colors of light emitted by bioluminescent beetles that use an identical substrate and chemiexcitation reaction sequence to generate light remain a challenging and controversial mechanistic conundrum. The crystal structures of two beetle luciferases with red- and blue-shifted light relative to the green yellow light of the common firefly species provide direct insight into the molecular origin of the bioluminescence color. The structure of a blue-shifted green-emitting luciferase from the firefly Amydetes vivianii is monomeric with a structural fold similar to the previously reported firefly luciferases. The only known naturally red-emitting luciferase from the glow-worm Phrixothrix hirtus exists as tetramers and octamers. Structural and computational analyses reveal varying aperture between the two domains enclosing the active site. Mutagenesis analysis identified two conserved loops that contribute to the color of the emitted light. These results are expected to advance comparative computational studies into the conformational landscape of the luciferase reaction sequence.

scholarly journals Biochemically diverse CRISPR-Cas9 orthologs

2020 ◽  
Author(s):  
Giedrius Gasiunas ◽  
Joshua K. Young ◽  
Tautvydas Karvelis ◽  
Darius Kazlauskas ◽  
Tomas Urbaitis ◽  
...  
Keyword(s):  
Temperature Dependence ◽  
Genome Editing ◽  
Guide Rna ◽  
Protospacer Adjacent Motif ◽  
Programmable Nucleases ◽  
Computational Analyses ◽  
Sequence Requirements ◽  
Insight Into ◽  
Biochemical Diversity

ABSTRACTCRISPR-Cas9 nucleases are abundant in microbes. To explore this largely uncharacterized diversity, we applied cell-free biochemical screens to rapidly assess the protospacer adjacent motif (PAM) and guide RNA (gRNA) requirements of novel Cas9 proteins. This approach permitted the characterization of 79 Cas9 orthologs with at least 7 distinct classes of gRNAs and 50 different PAM sequence requirements. PAM recognition spanned the entire spectrum of T-, A-, C-, and G-rich nucleotides ranging from simple di-nucleotide recognition to complex sequence strings longer than 4. Computational analyses indicated that most of this diversity came from 4 groups of interrelated sequences providing new insight into Cas9 evolution and efforts to engineer PAM recognition. A subset of Cas9 orthologs were purified and their activities examined further exposing additional biochemical diversity. This constituted both narrow and broad ranges of temperature dependence, staggered-end DNA target cleavage, and a requirement for longer stretches of homology between gRNA and DNA target to function robustly. In all, the diverse collection of Cas9 orthologs presented here sheds light on Cas9 evolution and provides a rich source of PAM recognition and other potentially desirable properties that may be mined to expand the genome editing toolbox with new RNA-programmable nucleases.

scholarly journals Substrate Specificity for Human Histidine Methyltransferase SETD3

2021 ◽  
Author(s):  
Jordi Hintzen ◽  
Huida Ma ◽  
Hao Deng ◽  
Apolonia Witecka ◽  
Steffen B. Andersen ◽  
...  
Keyword(s):  
Free Energy ◽  
Energy Barrier ◽  
Human Biology ◽  
Quantum Mechanical ◽  
Enzyme Assays ◽  
Free Energy Barrier ◽  
Methyl Transfer ◽  
Energy Simulations ◽  
Computational Analyses ◽  
Insight Into

Histidine methyltransferase SETD3 plays an important role in human biology and diseases. Previously, we showed that SETD3 catalyzes N3-methylation of histidine 73 in β-actin (Kwiatkowski et al., 2018). Here we report integrated synthetic, biocatalytic, biostructural and computational analyses on human SETD3-catalyzed methylation of β-actin peptides possessing histidine and its structurally and chemically diverse mimics. Our enzyme assays supported by biostructural analyses demonstrate that SETD3 has a broader substrate scope beyond histidine, including N-nucleophiles on the aromatic and aliphatic side chains. Quantum mechanical/molecular mechanical (QM/MM) molecular dynamics and free-energy simulations provide insight into binding geometries and the free energy barrier for the enzymatic methyl transfer to histidine mimics, further supporting experimental data that histidine is the superior SETD3 substrate over its analogs. This work demonstrates that human SETD3 has a potential to catalyze efficient methylation of several histidine mimics, overall providing mechanistic, biocatalytic and functional insight into β-actin histidine methylation by SETD3.

Computational Analyses of Mechanically Induced Collagen Fiber Remodeling in the Aortic Heart Valve

2003 ◽  
Vol 125 (4) ◽  
pp. 549-557 ◽  
Author(s):  
Niels J. B. Driessen ◽  
Ralf A. Boerboom ◽  
Jacques M. Huyghe ◽  
Carlijn V. C. Bouten ◽  
Frank P. T. Baaijens
Keyword(s):  
Experimental Data ◽  
Loading Condition ◽  
Heart Valves ◽  
Collagen Fiber ◽  
Fe Model ◽  
Mechanical Stimuli ◽  
Aortic Heart Valve ◽  
Strong Resemblance ◽  
Computational Analyses ◽  
Insight Into

To optimize the mechanical properties and integrity of tissue-engineered aortic heart valves, it is necessary to gain insight into the effects of mechanical stimuli on the mechanical behavior of the tissue using mathematical models. In this study, a finite-element (FE) model is presented to relate changes in collagen fiber content and orientation to the mechanical loading condition within the engineered construct. We hypothesized that collagen fibers aligned with principal strain directions and that collagen content increased with the fiber stretch. The results indicate that the computed preferred fiber directions run from commissure to commissure and show a strong resemblance to experimental data from native aortic heart valves.

scholarly journals Structural basis for channel conduction in the pump-like channelrhodopsin ChRmine

2021 ◽  
Author(s):  
Koichiro E. Kishi ◽  
Yoon Seok Kim ◽  
Masahiro Fukuda ◽  
Tsukasa Kusakizako ◽  
Elina Thadhani ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Transmembrane Helix ◽  
Action Spectrum ◽  
Light Sensitivity ◽  
Structural Basis ◽  
Ion Pump ◽  
Cryo Electron Microscopy ◽  
Architectural Features ◽  
Computational Analyses ◽  
Insight Into

ChRmine, a recently-discovered bacteriorhodopsin-like cation-conducting channelrhodopsin, exhibits puzzling properties (unusually-large photocurrents, exceptional red-shift in action spectrum, and extreme light-sensitivity) that have opened up new opportunities in optogenetics. ChRmine and its homologs function as light-gated ion channels, but by primary sequence more closely resemble ion pump rhodopsins; the molecular mechanisms for passive channel conduction in this family of proteins, as well as the unusual properties of ChRmine itself, have remained mysterious. Here we present the cryo-electron microscopy structure of ChRmine at 2.0 Å resolution. The structure reveals striking architectural features never seen before in channelrhodopsins including trimeric assembly, a short transmembrane-helix 3 unwound in the middle of the membrane, a prominently-twisting extracellular-loop 1, remarkably-large intracellular cavities and extracellular vestibule, and an unprecedented hydrophilic pore that extends through the center of the trimer, separate from the three individual monomer pores. Electrophysiological, spectroscopic, and computational analyses provide insight into conduction and gating of light-gated channels with these distinct design features, and point the way toward structure-guided creation of novel channelrhodopsins for optogenetic applications in biology.

scholarly journals The Ccr4-Not complex monitors the translating ribosome for codon optimality

2019 ◽  
Author(s):  
Robert Buschauer ◽  
Yoshitaka Matsuo ◽  
Ying-Hsin Chen ◽  
Najwa Alhusaini ◽  
Thomas Sweet ◽  
...  
Keyword(s):  
Mrna Stability ◽  
Mrna Decay ◽  
Specific Interaction ◽  
Ribosome Profiling ◽  
Translation Elongation ◽  
Cryo Electron Microscopy ◽  
A Site ◽  
Sense Codon ◽  
Decoding Efficiency ◽  
Insight Into

Control of mRNA decay rate is intimately connected to translation elongation but the spatial coordination of these events is poorly understood. The Ccr4-Not complex initiates mRNA decay through deadenylation and activation of decapping. Using a combination of cryo-electron microscopy, ribosome profiling and mRNA stability assays we show recruitment of Ccr4-Not to the ribosome via specific interaction of the Not5 subunit with the ribosomal E-site. This interaction only occurs when the ribosome lacks accommodated A-site tRNA, indicative of low codon optimality. Loss of Not5 results in the inability of the mRNA degradation machinery to sense codon optimality. Our analysis elucidates a physical link between the Ccr4-Not complex and the ribosome providing mechanistic insight into the coupling of decoding efficiency with mRNA stability.

Computational Analyses of Cavitating Control Elements in Cryogenic Environments

Volume 3 ◽  
2004 ◽  
Author(s):  
V. Ahuja ◽  
A. Hosangadi ◽  
J. Shipman
Keyword(s):  
Fluid Transport ◽  
Rocket Engine ◽  
Control Valves ◽  
Component Testing ◽  
Flow Metering ◽  
Thermodynamic Effects ◽  
Multi Phase ◽  
Computational Analyses ◽  
Phase Fluid ◽  
Insight Into

The safe and reliable operation of high pressure test stands for rocket engine and component testing places an increased emphasis on the performance of control valves and flow metering devices. In this paper, we present high fidelity computational analyses of cavitating venturi-type cryogenic control valves used to support rocket engine and component testing. The computational analyses are carried out with a generalized multi-phase formulation for cavitation in fluids operating in regimes where thermodynamic effects become important. The thermal effects and the accompanying property variations due to phase change are modeled rigorously. Thermal equilibrium is assumed and fluid thermodynamic properties are specified along the saturation line using the NIST-12 databank. The thermodynamic cavitation framework has been validated against experimental data of Hord [1] for hydrofoils operating in liquid nitrogen and liquid hydrogen. In this paper, we will discuss performance losses related to cryogenic control valves and provide insight into the physics of the dominant multi-phase fluid transport phenomena that are responsible for the choking like behavior of cryogenic control elements.

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