scholarly journals mCSM-membrane: predicting the effects of mutations on transmembrane proteins

2020 ◽  
Vol 48 (W1) ◽  
pp. W147-W153 ◽  
Author(s):  
Douglas E V Pires ◽  
Carlos H M Rodrigues ◽  
David B Ascher
Keyword(s):  
Membrane Proteins ◽  
Single Point ◽  
Point Mutations ◽  
Web Server ◽  
Protein Coding ◽  
Pathogenic Variants ◽  
Model Protein ◽  
Matthew’S Correlation Coefficient ◽  
Membrane Protein Stability ◽  
User Friendly

Abstract Significant efforts have been invested into understanding and predicting the molecular consequences of mutations in protein coding regions, however nearly all approaches have been developed using globular, soluble proteins. These methods have been shown to poorly translate to studying the effects of mutations in membrane proteins. To fill this gap, here we report, mCSM-membrane, a user-friendly web server that can be used to analyse the impacts of mutations on membrane protein stability and the likelihood of them being disease associated. mCSM-membrane derives from our well-established mutation modelling approach that uses graph-based signatures to model protein geometry and physicochemical properties for supervised learning. Our stability predictor achieved correlations of up to 0.72 and 0.67 (on cross validation and blind tests, respectively), while our pathogenicity predictor achieved a Matthew's Correlation Coefficient (MCC) of up to 0.77 and 0.73, outperforming previously described methods in both predicting changes in stability and in identifying pathogenic variants. mCSM-membrane will be an invaluable and dedicated resource for investigating the effects of single-point mutations on membrane proteins through a freely available, user friendly web server at http://biosig.unimelb.edu.au/mcsm_membrane.

scholarly journals Synthetic Genetic Codes Designed to Hinder Evolution

2019 ◽  
Author(s):  
Jonathan Calles ◽  
Isaac Justice ◽  
Detravious Brinkley ◽  
Alexa Garcia ◽  
Drew Endy
Keyword(s):  
Amino Acids ◽  
Single Point ◽  
Point Mutations ◽  
Translation System ◽  
Spontaneous Mutations ◽  
Protein Coding ◽  
Coding Sequences ◽  
Standard Code ◽  
Fail Safe

ABSTRACTOne challenge in engineering organisms is guaranteeing system behavior over many generations. Spontaneous mutations that arise before or during use can impact heterologous genetic functions, disrupt system integration, or change organism phenotype. Here, we propose restructuring the genetic code itself such that all point mutations in protein-coding sequences are selected against. Synthetic genetic systems so-encoded should “fail safely” in response to many individual spontaneous mutations. We designed a family of such fail-safe codes and analyzed their expected effect on the evolution of engineered organisms via simulation. We predict that fail-safe codes supporting expression of 20 or 15 amino acids could slow the evolution of proteins in so-encoded organisms to 30% or 0% the rate of standard-code organisms, respectively. We also designed quadruplet-codon codes that should be capable of encoding at least 20 amino acids while ensuring all single point mutations in protein-coding sequences are selected against. We show by in vitro experiments that a reduced set of 21 tRNA is capable of expressing a protein whose coding sequence is recoded to use a fail-safe code, whereas a standard-code encoding is not expressed. Our work suggests that a rationally depleted but otherwise natural translation system should yield biological systems with intrinsically reduced evolutionary capacity, and that so-encoded hypoevolvable organisms might be less likely to invade new niches or outcompete native populations.

scholarly journals MTR-Viewer: identifying regions within genes under purifying selection

2019 ◽  
Vol 47 (W1) ◽  
pp. W121-W126 ◽  
Author(s):  
Michael Silk ◽  
Slavé Petrovski ◽  
David B Ascher
Keyword(s):  
Selective Pressure ◽  
Web Server ◽  
Purifying Selection ◽  
Genome Sequences ◽  
Protein Coding ◽  
Sliding Windows ◽  
Alternative Transcripts ◽  
Missense Variation ◽  
User Friendly ◽  
Tolerance Ratio

Abstract Advances in genomic sequencing have enormous potential to revolutionize personalized medicine, however distinguishing disease-causing from benign variants remains a challenge. The increasing number of human genome and exome sequences available has revealed areas where unfavourable variation is removed through purifying selection. Here, we present the MTR-Viewer, a web-server enabling easy visualization at the gene or variant level of the Missense Tolerance Ratio (MTR), a measure of regional intolerance to missense variation calculated using variation from 240 000 exome and genome sequences. The MTR-Viewer enables exploration of MTR calculations, using different sliding windows, for over 18 000 human protein-coding genes and 85 000 alternative transcripts. Users can also view MTR scores calculated for specific ethnicities, to enable easy exploration of regions that may be under different selective pressure. The spatial distribution of population and known disease variants is also displayed on the protein's domain structure. Intolerant regions were found to be highly enriched for ClinVar pathogenic and COSMIC somatic missense variants (Mann–Whitney U test P < 2.2 × 10−16). As the MTR is not biased by known domains and protein features, it can highlight functionally important regions within genes overlooked or inaccessible by traditional methods. MTR-Viewer is freely available via a user friendly web-server at http://biosig.unimelb.edu.au/mtr-viewer/.

scholarly journals Fail-safe genetic codes designed to intrinsically contain engineered organisms

2019 ◽  
Vol 47 (19) ◽  
pp. 10439-10451 ◽  
Author(s):  
Jonathan Calles ◽  
Isaac Justice ◽  
Detravious Brinkley ◽  
Alexa Garcia ◽  
Drew Endy
Keyword(s):  
Amino Acids ◽  
System Integration ◽  
Single Point ◽  
Point Mutations ◽  
Spontaneous Mutations ◽  
Protein Coding ◽  
Coding Sequences ◽  
Encoded Proteins ◽  
Fail Safe ◽  
Translation Systems

Abstract One challenge in engineering organisms is taking responsibility for their behavior over many generations. Spontaneous mutations arising before or during use can impact heterologous genetic functions, disrupt system integration, or change organism phenotype. Here, we propose restructuring the genetic code itself such that point mutations in protein-coding sequences are selected against. Synthetic genetic systems so-encoded should fail more safely in response to most spontaneous mutations. We designed fail-safe codes and simulated their expected effects on the evolution of so-encoded proteins. We predict fail-safe codes supporting expression of 20 or 15 amino acids could slow protein evolution to ∼30% or 0% the rate of standard-encoded proteins, respectively. We also designed quadruplet-codon codes that should ensure all single point mutations in protein-coding sequences are selected against while maintaining expression of 20 or more amino acids. We demonstrate experimentally that a reduced set of 21 tRNAs is capable of expressing a protein encoded by only 20 sense codons, whereas a standard 64-codon encoding is not expressed. Our work suggests that biological systems using rationally depleted but otherwise natural translation systems should evolve more slowly and that such hypoevolvable organisms may be less likely to invade new niches or outcompete native populations.

Cupin Variants as Macromolecular Ligand Library for Stereoselective Michael Addition of Nitroalkanes

2019 ◽  
Author(s):  
Nobutaka Fujieda ◽  
Miho Yuasa ◽  
Yosuke Nishikawa ◽  
Genji Kurisu ◽  
Shinobu Itoh ◽  
...  
Keyword(s):  
Michael Addition ◽  
In Silico ◽  
Addition Reaction ◽  
Single Point ◽  
Point Mutations ◽  
Unsaturated Ketone ◽  
Michael Addition Reaction ◽  
Beta Barrel ◽  
Cupin Superfamily ◽  
Single Point Mutations

Cupin superfamily proteins (TM1459) work as a macromolecular ligand framework with a double-stranded beta-barrel structure ligating to a Cu ion through histidine side chains. Variegating the first coordination sphere of TM1459 revealed that H52A and H54A/H58A mutants effectively catalyzed the diastereo- and enantio-selective Michael addition reaction of nitroalkanes to an α,β-unsaturated ketone. Moreover, in silico substrate docking signified C106N and F104W single-point mutations, which inverted the diastereoselectivity of H52A and further improved the stereoselectivity of H54A/H58A, respectively.

PlncRNADB: A Repository of Plant lncRNAs and lncRNA-RBP Protein Interactions

2019 ◽  
Vol 14 (7) ◽  
pp. 621-627 ◽  
Author(s):  
Youhuang Bai ◽  
Xiaozhuan Dai ◽  
Tiantian Ye ◽  
Peijing Zhang ◽  
Xu Yan ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Populus Trichocarpa ◽  
Noncoding Rnas ◽  
Reference Database ◽  
Protein Coding ◽  
Arabidopsis Lyrata ◽  
User Friendly

Background: Long noncoding RNAs (lncRNAs) are endogenous noncoding RNAs, arbitrarily longer than 200 nucleotides, that play critical roles in diverse biological processes. LncRNAs exist in different genomes ranging from animals to plants. Objective: PlncRNADB is a searchable database of lncRNA sequences and annotation in plants. Methods: We built a pipeline for lncRNA prediction in plants, providing a convenient utility for users to quickly distinguish potential noncoding RNAs from protein-coding transcripts. Results: More than five thousand lncRNAs are collected from four plant species (Arabidopsis thaliana, Arabidopsis lyrata, Populus trichocarpa and Zea mays) in PlncRNADB. Moreover, our database provides the relationship between lncRNAs and various RNA-binding proteins (RBPs), which can be displayed through a user-friendly web interface. Conclusion: PlncRNADB can serve as a reference database to investigate the lncRNAs and their interaction with RNA-binding proteins in plants. The PlncRNADB is freely available at http://bis.zju.edu.cn/PlncRNADB/.

Single-Point Mutations in Qβ Virus-like Particles Change Binding to Cells

2021 ◽  
Author(s):  
Marisa L. Martino ◽  
Stephen N. Crooke ◽  
Marianne Manchester ◽  
M.G. Finn
Keyword(s):  
Single Point ◽  
Point Mutations ◽  
Virus Like Particles ◽  
Single Point Mutations

scholarly journals Mutational patterns and clonal evolution from diagnosis to relapse in pediatric acute lymphoblastic leukemia

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Shumaila Sayyab ◽  
Anders Lundmark ◽  
Malin Larsson ◽  
Markus Ringnér ◽  
Sara Nystedt ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Large Scale ◽  
Somatic Mutations ◽  
Lymphoblastic Leukemia ◽  
Clonal Evolution ◽  
Point Mutations ◽  
Driver Genes ◽  
Protein Coding ◽  
Pediatric Acute Lymphoblastic Leukemia ◽  
Evolutionary Trajectories

AbstractThe mechanisms driving clonal heterogeneity and evolution in relapsed pediatric acute lymphoblastic leukemia (ALL) are not fully understood. We performed whole genome sequencing of samples collected at diagnosis, relapse(s) and remission from 29 Nordic patients. Somatic point mutations and large-scale structural variants were called using individually matched remission samples as controls, and allelic expression of the mutations was assessed in ALL cells using RNA-sequencing. We observed an increased burden of somatic mutations at relapse, compared to diagnosis, and at second relapse compared to first relapse. In addition to 29 known ALL driver genes, of which nine genes carried recurrent protein-coding mutations in our sample set, we identified putative non-protein coding mutations in regulatory regions of seven additional genes that have not previously been described in ALL. Cluster analysis of hundreds of somatic mutations per sample revealed three distinct evolutionary trajectories during ALL progression from diagnosis to relapse. The evolutionary trajectories provide insight into the mutational mechanisms leading relapse in ALL and could offer biomarkers for improved risk prediction in individual patients.

scholarly journals Membrane Protein Stabilization Strategies for Structural and Functional Studies

Membranes ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 155
Author(s):  
Ekaitz Errasti-Murugarren ◽  
Paola Bartoccioni ◽  
Manuel Palacín
Keyword(s):  
Membrane Proteins ◽  
Membrane Protein ◽  
Lipid Membrane ◽  
Protein Stabilization ◽  
New Drugs ◽  
Cell Physiology ◽  
Protein Preparation ◽  
Functional Studies ◽  
Membrane Protein Stability ◽  
Druggable Targets

Accounting for nearly two-thirds of known druggable targets, membrane proteins are highly relevant for cell physiology and pharmacology. In this regard, the structural determination of pharmacologically relevant targets would facilitate the intelligent design of new drugs. The structural biology of membrane proteins is a field experiencing significant growth as a result of the development of new strategies for structure determination. However, membrane protein preparation for structural studies continues to be a limiting step in many cases due to the inherent instability of these molecules in non-native membrane environments. This review describes the approaches that have been developed to improve membrane protein stability. Membrane protein mutagenesis, detergent selection, lipid membrane mimics, antibodies, and ligands are described in this review as approaches to facilitate the production of purified and stable membrane proteins of interest for structural and functional studies.

scholarly journals In Vivo Evidence for TonB Dimerization

2003 ◽  
Vol 185 (19) ◽  
pp. 5747-5754 ◽  
Author(s):  
Annette Sauter ◽  
S. Peter Howard ◽  
Volkmar Braun
Keyword(s):  
Single Point ◽  
Point Mutations ◽  
Ferric Citrate ◽  
Site Directed Mutagenesis ◽  
Toxin Gene ◽  
Transmembrane Region ◽  
Dimer Formation ◽  
Citrate Transport ◽  
Hybrid Proteins

ABSTRACT TonB, in complex with ExbB and ExbD, is required for the energy-dependent transport of ferric siderophores across the outer membrane of Escherichia coli, the killing of cells by group B colicins, and infection by phages T1 and φ80. To gain insights into the protein complex, TonB dimerization was studied by constructing hybrid proteins from complete TonB (containing amino acids 1 to 239) [TonB(1-239)] and the cytoplasmic fragment of ToxR which, when dimerized, activates the transcription of the cholera toxin gene ctx. ToxR(1-182)-TonB(1-239) activated the transcription of lacZ under the control of the ctx promoter (P ctx ::lacZ). Replacement of the TonB transmembrane region by the ToxR transmembrane region resulted in the hybrid proteins ToxR(1-210)-TonB(33-239) and ToxR(1-210)-TonB(164-239), of which only the latter activated P ctx ::lacZ transcription. Dimer formation was reduced but not abolished in a mutant lacking ExbB and ExbD, suggesting that these complex components may influence dimerization but are not strictly required and that the N-terminal cytoplasmic membrane anchor and the C-terminal region are important for dimer formation. The periplasmic TonB fragment, TonB(33-239), inhibits ferrichrome and ferric citrate transport and induction of the ferric citrate transport system. This competition provided a means to positively screen for TonB(33-239) mutants which displayed no inhibition. Single point mutations of inactive fragments selected in this manner were introduced into complete TonB, and the phenotypes of the TonB mutant strains were determined. The mutations located in the C-terminal half of TonB, three of which (Y163C, V188E, and R204C) were obtained separately by site-directed mutagenesis, as was the isolated F230V mutation, were studied in more detail. They displayed different activity levels for various TonB-dependent functions, suggesting function-related specificities which reflect differences in the interactions of TonB with various transporters and receptors.

MinD directly interacting with FtsZ at the H10 helix suggests a model for robust activation of MinC to destabilize FtsZ polymers

2017 ◽  
Vol 474 (18) ◽  
pp. 3189-3205 ◽  
Author(s):  
Ashoka Chary Taviti ◽  
Tushar Kant Beuria
Keyword(s):  
Cell Division ◽  
Single Point ◽  
Point Mutations ◽  
Direct Interaction ◽  
Ring Formation ◽  
Z Ring ◽  
Single Point Mutations ◽  
Biochemical Analyses ◽  
Ftsz Assembly ◽  
The Mind

Cell division in bacteria is a highly controlled and regulated process. FtsZ, a bacterial cytoskeletal protein, forms a ring-like structure known as the Z-ring and recruits more than a dozen other cell division proteins. The Min system oscillates between the poles and inhibits the Z-ring formation at the poles by perturbing FtsZ assembly. This leads to an increase in the FtsZ concentration at the mid-cell and helps in Z-ring positioning. MinC, the effector protein, interferes with Z-ring formation through two different mechanisms mediated by its two domains with the help of MinD. However, the mechanism by which MinD triggers MinC activity is not yet known. We showed that MinD directly interacts with FtsZ with an affinity stronger than the reported MinC–FtsZ interaction. We determined the MinD-binding site of FtsZ using computational, mutational and biochemical analyses. Our study showed that MinD binds to the H10 helix of FtsZ. Single-point mutations at the charged residues in the H10 helix resulted in a decrease in the FtsZ affinity towards MinD. Based on our findings, we propose a novel model for MinCD–FtsZ interaction, where MinD through its direct interaction with FtsZ would trigger MinC activity to inhibit FtsZ functions.

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