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51
CUPSAT: prediction of protein stability upon point mutations
- Nucleic Acids Res
, 2006
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Protein thermostability calculations using alchemical free energy simulations
- Biophys. J
, 2010
"... ABSTRACT Thermal stability of proteins is crucial for both biotechnological and therapeutic applications. Rational protein engineering therefore frequently aims at increasing thermal stability by introducing stabilizing mutations. The accurate prediction of the thermodynamic consequences caused by m ..."
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ABSTRACT Thermal stability of proteins is crucial for both biotechnological and therapeutic applications. Rational protein engineering therefore frequently aims at increasing thermal stability by introducing stabilizing mutations. The accurate prediction of the thermodynamic consequences caused by mutations, however, is highly challenging as thermal stability changes are caused by alterations in the free energy of folding. Growing computational power, however, increasingly allows us to use alchemical free energy simulations, such as free energy perturbation or thermodynamic integration, to calculate free energy differences with relatively high accuracy. In this article, we present an automated protocol for setting up alchemical free energy calculations for mutations of naturally occurring amino acids (except for proline) that allows an unprecedented, automated screening of large mutant libraries. To validate the developed protocol, we calculated thermodynamic stability differences for 109 mutations in the microbial Ribonuclease Barnase. The obtained quantitative agreement with experimental data illustrates the potential of the approach in protein engineering and design.
Bioinformatics for personal genome interpretation. Briefings in Bioinformatics
, 2012
"... Abstract An international consortium released the first draft sequence of the human genome 10 years ago. Although the analysis of this data has suggested the genetic underpinnings of many diseases, we have not yet been able to fully quantify the relationship between genotype and phenotype. Thus, a ..."
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Abstract An international consortium released the first draft sequence of the human genome 10 years ago. Although the analysis of this data has suggested the genetic underpinnings of many diseases, we have not yet been able to fully quantify the relationship between genotype and phenotype. Thus, a major current effort of the scientific community focuses on evaluating individual predispositions to specific phenotypic traits given their genetic backgrounds. Many resources aim to identify and annotate the specific genes responsible for the observed phenotypes. Some of these use intra-species genetic variability as a means for better understanding this relationship. In addition, several online resources are now dedicated to collecting single nucleotide variants and other types of variants, and annotating their functional effects and associations with phenotypic traits. This information has enabled researchers to develop bioinformatics tools to analyze the rapidly increasing amount of newly extracted variation data and to predict the effect of uncharacterized variants. In this work, we review the most important developments in the fieldçthe databases and bioinformatics tools that will be of utmost importance in our concerted effort to interpret the human variome.
Crohn's disease risk alleles on the NOD2 locus have been maintained by natural selection on standing variation. Mol Biol Evol 29: 1569–1585. doi: 10.1093/molbev/mss006 PMID: 22319155
, 2012
"... Risk alleles for complex diseases are widely spread throughout human populations. However, little is known about the geographic distribution and frequencies of risk alleles, which may contribute to differences in disease susceptibility and prevalence among populations. Here, we focus on Crohn’s dise ..."
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Risk alleles for complex diseases are widely spread throughout human populations. However, little is known about the geographic distribution and frequencies of risk alleles, which may contribute to differences in disease susceptibility and prevalence among populations. Here, we focus on Crohn’s disease (CD) as a model for the evolutionary study of complex disease alleles. Recent genome-wide association studies and classical linkage analyses have identified more than 70 susceptible genomic regions for CD in Europeans, but only a few have been confirmed in non-European populations. Our analysis of eight European-specific susceptibility genes using HapMap data shows that at the NOD2 locus the CD-risk alleles are linked with a haplotype specific to CEU at a frequency that is significantly higher compared with the entire genome. We subsequently examined nine global populations and found that the CD-risk alleles spread through hitchhiking
www.mdpi.com/journal/ijms Improvement of Thermal Stability via Outer-Loop Ion Pair Interaction of Mutated T1 Lipase from Geobacillus zalihae
, 2012
"... Abstract: Mutant D311E and K344R were constructed using site-directed mutagenesis to introduce an additional ion pair at the inter-loop and the intra-loop, respectively, to determine the effect of ion pairs on the stability of T1 lipase isolated from Geobacillus zalihae. A series of purification ste ..."
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Abstract: Mutant D311E and K344R were constructed using site-directed mutagenesis to introduce an additional ion pair at the inter-loop and the intra-loop, respectively, to determine the effect of ion pairs on the stability of T1 lipase isolated from Geobacillus zalihae. A series of purification steps was applied, and the pure lipases of T1, D311E and K344R were obtained. The wild-type and mutant lipases were analyzed using circular dichroism. The Tm for T1 lipase, D311E lipase and K344R lipase were approximately 68.52 °C, 70.59 °C and 68.54 °C, respectively. Mutation at D311 increases the stability of T1 lipase and exhibited higher Tm as compared to the wild-type and K344R. Based on the above, D311E lipase was chosen for further study. D311E lipase was successfully crystallized using the sitting drop vapor diffusion method. The crystal was diffracted at 2.1 Å using an in-house X-ray beam and belonged to the monoclinic space group C2 with the unit cell parameters a = 117.32 Å, b = 81.16 Å and c = 100.14 Å. Structural analysis showed the existence of an additional ion pair around E311 in the structure of D311E. TheInt. J. Mol. Sci. 2012, 13 944 additional ion pair in D311E may regulate the stability of this mutant lipase at high
An Evolutionary Analysis of Antigen Processing and Presentation across Different Timescales Reveals Pervasive Selection
"... The antigenic repertoire presented by MHC molecules is generated by the antigen processing and presentation (APP) pathway. We analyzed the evolutionary history of 45 genes involved in APP at the inter- and intra-species level. Results showed that 11 genes evolved adaptively in mammals. Several posit ..."
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The antigenic repertoire presented by MHC molecules is generated by the antigen processing and presentation (APP) pathway. We analyzed the evolutionary history of 45 genes involved in APP at the inter- and intra-species level. Results showed that 11 genes evolved adaptively in mammals. Several positively selected sites involve positions of fundamental importance to the protein function (e.g. the TAP1 peptide-binding domains, the sugar binding interface of langerin, and the CD1D trafficking signal region). In CYBB, all selected sites cluster in two loops protruding into the endosomal lumen; analysis of missense mutations responsible for chronic granulomatous disease (CGD) showed the action of different selective forces on the very same gene region, as most CGD substitutions involve aminoacid positions that are conserved in all mammals. As for ERAP2, different computational methods indicated that positive selection has driven the recurrent appearance of protein-destabilizing variants during mammalian evolution. Application of a population-genetics phylogenetics approach showed that purifying selection represented a major force acting on some APP components (e.g. immunoproteasome subunits and chaperones) and allowed identification of positive selection events in the human lineage. We also investigated the evolutionary history of APP genes in human populations by developing a new approach that uses several
Point mutations in protein globular domains: contributions from function, stability and misfolding
- J Mol Biol
, 2006
"... Several contrasting hypotheses have been formulated about the influence of functional and conformational properties, like stability and avoidance of misfolding, on the evolution of protein globular domains. Selection at functional sites has been suggested to be detrimental to stability or coupled t ..."
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Several contrasting hypotheses have been formulated about the influence of functional and conformational properties, like stability and avoidance of misfolding, on the evolution of protein globular domains. Selection at functional sites has been suggested to be detrimental to stability or coupled to it. Avoidance of misfolding may be achieved by discarding misfoldingprone sequences or by maintaining a stable native state and thus destabilizing partially or fully unfolded states from which misfolding can take place. We have performed a hierarchical analysis of a large database of point mutations to dissect the relative contributions of function, stability and misfolding in the evolution of natural sequences. We show that at catalytic sites, selection for function overrules selection for stability but find no evidence for an anticorrelation between function and stability. Selection for stability plays a secondary role at binding sites, but is not fully coupled to selection for function. Remarkably, we did not find a selective pressure against misfolding-prone sequences in globular proteins at the level of individual positions. We suggest that such a selection would compromise native-state stability due to a correlation between the stabilities of native and misfolded states. Stabilization of the native state is the most frequent way in which natural proteins avoid misfolding.
RosettaBackrub—a web server for flexible backbone protein structure modeling
, 2010
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Review Computational and Experimental Approaches to Reveal the Effects of Single Nucleotide Polymorphisms with Respect to Disease Diagnostics
, 2014
"... Abstract: DNA mutations are the cause of many human diseases and they are the reason for natural differences among individuals by affecting the structure, function, interactions, and other properties of DNA and expressed proteins. The ability to predict whether a given mutation is disease-causing or ..."
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Abstract: DNA mutations are the cause of many human diseases and they are the reason for natural differences among individuals by affecting the structure, function, interactions, and other properties of DNA and expressed proteins. The ability to predict whether a given mutation is disease-causing or harmless is of great importance for the early detection of patients with a high risk of developing a particular disease and would pave the way for personalized medicine and diagnostics. Here we review existing methods and techniques to study and predict the effects of DNA mutations from three different perspectives: in silico, in vitro and in vivo. It is emphasized that the problem is complicated and successful detection of a pathogenic mutation frequently requires a combination of several methods and a knowledge of the biological phenomena associated with the corresponding macromolecules.
KD4v: Comprehensible Knowledge Discovery System for Missense Variant
- Nucleic Acids Res
, 2012
"... A major challenge in the post-genomic era is a better understanding of how human genetic alter-ations involved in disease affect the gene products. The KD4v (Comprehensible Knowledge Discovery System for Missense Variant) server allows to char-acterize and predict the phenotypic effects (deleteri-ou ..."
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A major challenge in the post-genomic era is a better understanding of how human genetic alter-ations involved in disease affect the gene products. The KD4v (Comprehensible Knowledge Discovery System for Missense Variant) server allows to char-acterize and predict the phenotypic effects (deleteri-ous/neutral) of missense variants. The server provides a set of rules learned by Induction Logic Programming (ILP) on a set of missense variants described by conservation, physico-chemical, func-tional and 3D structure predicates. These rules are interpretable by non-expert humans and are used to accurately predict the deleterious/neutral status of an unknown mutation. The web server is available at
