scholarly journals Application of computational algorithms to assess the functionality of non-synonymous substitutions in MHC DRB gene of Nigerian goats

Genetika ◽  
2017 ◽  
Vol 49 (1) ◽  
pp. 63-76
Author(s):  
Abdulmojeed Yakubu ◽  
Adebowale Salako ◽  
Donato de ◽  
Ikhide Imumorin
Keyword(s):  
Amino Acid ◽  
Mhc Class Ii ◽  
Protein Function ◽  
Gene Families ◽  
Acid Sites ◽  
Amino Acid Substitutions ◽  
Nucleotide Polymorphisms ◽  
Antigen Binding Site ◽  
Exon 2 ◽  
Drb Gene

The Major Histocompatibility Complex (MHC) contains highly variable multi-gene families, which play a key role in the adaptive immune response within vertebrates. Among the Capra MHC class II genes, the expressed DRB locus is highly polymorphic, particularly in exon 2, which encodes the antigen-binding site. Models of variable non-synonymous/synonymous rate ratios among sites may provide important insights into functional constraints at different amino acid sites and may be used to detect sites under positive selection. Many non-synonymous single nucleotide polymorphisms (nsSNPs) at the DRB locus in goats are suspected to impact protein function. This study, therefore, aimed at comparing the efficiency of six computational approaches to predict the likelihood of a particular non-synonymous (amino acid change) coding SNP to cause a functional impact on the protein. This involved the use of PANTHER, SNAP, SIFT, PolyPhen-2, PROVEAN and nsSNPAnalyzer bioinformatics analytical tools in detecting harmful and beneficial effects at H57G, Y89R, V104D and Y112I substitutions in the peptide binding region of the DRB gene of Nigerian goats. The results from PANTHER analysis revealed that H57G, Y89R and Y112I substitutions (Pdeleterious= 0.113, 0.204 and 0.472, respectively) were beneficial; while that of V104D was deleterious (Pdeleterious= 0.756), an indication that it was non-neutral. As regards the SNAP approach, H57G and Y89R substitutions were returned neutral with expected accuracy of 53 and 69%, respectively while V104D and Y112I substitutions were harmful. H57G and Y89R substitutions were also found harmless in the SIFT analysis. However, only H57G (PROVEAN) and V104D (nsSNPAnalyzer) amino acid substitutions were found to be beneficial. Interestingly, the predicted 3D structures of both native and mutant DRB protein appeared similar as validated by Ramachandran plots. The consensus reached by PANTHER, SNAP, SIFT and PolyPhen-2 approaches on the neutrality especially of H57G (PROVEAN inclusive) and Y89R amino acid substitutions may be used in search of disease resistant genotypes at the DRB locus of Nigerian goats.

A STRUCTURAL BIOINFORMATICS APPROACH TO THE ANALYSIS OF NONSYNONYMOUS SINGLE NUCLEOTIDE POLYMORPHISMS (nsSNPs) AND THEIR RELATION TO DISEASE

2007 ◽  
Vol 05 (06) ◽  
pp. 1297-1318 ◽  
Author(s):  
CATHERINE L. WORTH ◽  
G. RICHARD J. BICKERTON ◽  
ADRIAN SCHREYER ◽  
JULIA R. FORMAN ◽  
TAMMY M. K. CHENG ◽  
...  
Keyword(s):  
Amino Acid ◽  
Single Nucleotide Polymorphisms ◽  
Structure Prediction ◽  
Three Dimensional ◽  
Comparative Modeling ◽  
Divergent Evolution ◽  
Amino Acid Substitutions ◽  
Nucleotide Polymorphisms ◽  
Single Nucleotide ◽  
Human Proteins

The prediction of the effects of nonsynonymous single nucleotide polymorphisms (nsSNPs) on function depends critically on exploiting all information available on the three-dimensional structures of proteins. We describe software and databases for the analysis of nsSNPs that allow a user to move from SNP to sequence to structure to function. In both structure prediction and the analysis of the effects of nsSNPs, we exploit information about protein evolution, in particular, that derived from investigations on the relation of sequence to structure gained from the study of amino acid substitutions in divergent evolution. The techniques developed in our laboratory have allowed fast and automated sequence-structure homology recognition to identify templates and to perform comparative modeling; as well as simple, robust, and generally applicable algorithms to assess the likely impact of amino acid substitutions on structure and interactions. We describe our strategy for approaching the relationship between SNPs and disease, and the results of benchmarking our approach — human proteins of known structure and recognized mutation.

Single Nucleotide Polymorphisms Influence the Phenotype of Mutations P618A and R1336W found in the ADAMTS13 Gene of Congenital TTP Patients.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2977-2977
Author(s):  
Barbara Plaimauer ◽  
Gabriele Mohr ◽  
Waltraud Wernhart ◽  
Katharina Bruno ◽  
Gerhard Antoine ◽  
...  
Keyword(s):  
Amino Acid ◽  
Single Nucleotide Polymorphisms ◽  
Single Amino Acid ◽  
Paternal Allele ◽  
Amino Acid Substitutions ◽  
Common Snps ◽  
Nucleotide Polymorphisms ◽  
Adamts13 Activity ◽  
Maternal Allele ◽  
Single Nucleotide

Abstract ADAMTS13 cleaves plasmatic von Willebrand factor (VWF) between Tyr1605 and Met1606 and regulates thereby the hemostatic activity of VWF. Mutations in the ADAMTS13 gene leading to severe ADAMTS13 deficiency have been found in patients with congenital thrombotic thrombocytopenic purpura (TTP). We have analyzed the ADAMTS13 gene defects in two brothers with hereditary TTP [Antoine et al, Brit. J. Hematol., 2003] where we detected a total of six nucleotide exchanges causing point mutations. On the maternal allele we found an accumulation of five amino acid substitutions (R7W, Q448E, P618A, A732V, R1336W) and on the paternal allele a stop mutation (Q44X) leading to premature protein termination in the propeptide region. Both brothers were double heterozygotes with < 3% of ADAMTS13 activity, whereas their asymptomatic parents have ~ 50% activity. Four (R7W, Q448E, P618A, A732V) of the five maternal mutations constitute single nucleotide polymorphisms (SNP) but R1336W was identified as novel rare mutation in the second cub domain. To evaluate the biologic phenotype of a given haplotype, e.g. the functional significance of the presence of the various SNPs, we analyzed the functional impact of the individual mutations on ADAMTS13 antigen levels and ADAMTS13 activity. A series of mutant ADAMTS13 molecules was expressed which contained either single amino acid substitutions or combinations of mutations with each other. We found that the common SNPs R7W, Q448E and A732V, as single mutations, had either no or only a minor impact on ADAMTS13 secretion and ADAMTS13 activity, whereas P618A and R1336W conferred a dominant adverse effect on ADAMTS13 secretion levels. Co-expression of SNPs R7W or Q448E with SNP P618A lead to improved ADAMTS13 secretion levels and could therefore partly attenuate the detrimental effect of P618A. Concomitant expression of all four SNPs reconstituted secretion levels similar to wild-type implicating a complex synergistically interaction of SNPs located in different ADAMTS13 domain regions, however, functional activity was impaired to 50%. Mutation R1336W was shown to be, as a single amino acid exchange, responsible for reduced ADAMTS13 antigen levels, but in contrast to P618A, the negative effect of R1336W was rather enhanced by the co-expression of R7W and Q448E, than rescued, leading to the total absence of ADAMTS13 secretion from the maternal allele. Our findings provide for the first time evidence that fairly common SNPs, dependent on the presence or absence of other mutations, may differently modulate functional ADAMTS13 protease levels.

scholarly journals Understanding the origins of loss of protein function by analyzing the effects of thousands of variants on activity and abundance

2020 ◽  
Author(s):  
Matteo Cagiada ◽  
Kristoffer E. Johansson ◽  
Audronė Valančiūtė ◽  
Sofie V. Nielsen ◽  
Rasmus Hartmann-Petersen ◽  
...  
Keyword(s):  
Experimental Data ◽  
Amino Acid ◽  
Enzymatic Activity ◽  
Indirect Effects ◽  
Protein Function ◽  
Evolutionary Conservation ◽  
Amino Acid Substitutions ◽  
Loss Of Function ◽  
Basic Understanding ◽  
Computational Analyses

AbstractUnderstanding and predicting how amino acid substitutions affect proteins is key to practical uses of proteins, and to our basic understanding of protein function and evolution. Amino acid changes may affect protein function in a number of ways including direct perturbations of activity or indirect effects on protein folding and stability. We have analysed 6749 experimentally determined variant effects from multiplexed assays on abundance and activity in two proteins (NUDT15 and PTEN) to quantify these effects, and find that a third of the variants cause loss of function, and about half of loss-of-function variants also have low cellular abundance. We analyse the structural and mechanistic origins of loss of function, and use the experimental data to find residues important for enzymatic activity. We performed computational analyses of protein stability and evolutionary conservation and show how we may predict positions where variants cause loss of activity or abundance.

Reduced Immunogenicity of FVIII Peptides by Rational Modifications of An Immunodominant T-Cell Epitope.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 220-220
Author(s):  
Ruth A. Ettinger ◽  
Joseph A. Liberman ◽  
Douglas C. Bolgiano ◽  
Arthur R. Thompson ◽  
Kathleen P. Pratt
Keyword(s):  
T Cells ◽  
Amino Acid ◽  
T Cell ◽  
Mhc Class Ii ◽  
Hemophilia A ◽  
Peptide Binding ◽  
Cell Epitope ◽  
Class Ii ◽  
Amino Acid Substitutions ◽  
T Cell Epitope

Abstract Abstract 220 Introduction. The development of antibodies that interfere with factor VIII (FVIII) pro-coagulant activity, often referred to as “inhibitors”, can complicate the treatment of hemophilia A. These alloimmune responses, as well as the rare development of autoimmune FVIII inhibitors, are associated with significant morbidity and mortality. The production of anti-FVIII antibodies follows stimulation of helper T cells by epitopes in FVIII. An immunodominant HLA-DRB1*0101-restricted T-cell epitope was recognized by CD4+ T cells from a mild hemophilia A inhibitor subject and from his brother, who had a sub-clinical inhibitor (James et al., J Thromb Haemost 5: 2399-2407, 2007). Their CD4+ T cells recognized overlapping synthetic peptides with sequences corresponding to FVIII residues 2186-2205, 2187-2205 and 2194-2213. Nineteen T-cell clones recognizing this epitope were isolated, with phenotypes representing four distinct T-cell lineages. Aims: (1) to evaluate the promiscuity/immunodominance of an HLA-DRB1*0101-restricted T-cell epitope in FVIII; (2) to introduce amino acid substitutions that will prevent presentation of this epitope to the immune system by DR0101 and by other DR alleles. Methods. The minimal epitope and MHC Class II (DR0101) “anchor” residues were determined using a competition assay measuring displacement of a labeled peptide having high affinity for recombinant DR0101 by a series of FVIII peptides. Peptide concentrations at which 50% inhibition of the labeled peptide binding occurred (IC50) were obtained by regression analysis. Binding of the peptides to five additional DR alleles was evaluated directly using recombinant proteins; predicted binding of peptides to additional DR alleles was evaluated using the program ProPred. Proliferation and cytokine production by the clones in response to wild-type and modified peptides were measured, and the concentrations at which half-maximal T-cell responses (EC50) to the FVIII peptides occurred were determined. Results. Binding of truncated peptides to DR0101 identified FVIII2194-2205 as the minimal epitope. Binding of FVIII2194-2205 peptides with single Arg substitutions identified F2196, M2199, A2201 and S2204 as anchor residues at positions 1, 4, 6 and 9, respectively, corresponding to peptide-binding pockets seen in the crystal structure of a DR0101-peptide complex. The relative binding of Ala-substituted peptides confirmed that F2196 and M2199 are anchor residues. T-cell stimulation requires recognition of peptides by both the Class II receptor and the T-cell receptor (TCR). Sequences of TCR variable regions (TCRBVs) expressed by the clones were identified as TCRBV20-1*01 (3 VDJ combinations), TCRBV6-6*01, TCRBV5-1*01, and TCRBV6-1*01, indicating at least six different T-cell progenitors recognized this epitope. The clones were next stimulated with peptides having modified epitopes. Strikingly, none proliferated or secreted cytokines when stimulated by FVIII2194-2205, F2196A, which also showed an IC50 > 10 μM when tested for binding to DR0101, DR0301, DR0401, DR1101, DR1104, and DR1501. Substitutions at other anchor positions affected binding to some but not all of the DR proteins. Predicted binding of the F2196A variant to 51 DR alleles was analyzed using ProPred; none bound at a threshold stringency of 10% (low stringency, thus the predicted epitopes included those with lower calculated affinities). In preparation for directly testing the immunogenicity of additional substitutions, all possible amino acid substitutions at position 2196 were evaluated using ProPred. 13 of 19 possible substitutions were predicted to prevent FVIII2194-2205 binding to all 51 DR alleles included in the algorithm (with a 3% threshold = intermediate stringency). Conclusions. MHC class II anchor residues and TCR contact sites for an immunodominant HLA-DRB1*0101-restricted T-cell epitope have been mapped precisely. Both measured and predicted effects of amino acid substitutions indicated that this F2196 is essential for effective presentation of this epitope by multiple DR alleles. Effects of various sequence modifications on FVIII function, conformation and immunogenicity are currently being evaluated using recombinant FVIII and FVIII C2 domain proteins to indicate their possible therapeutic potential. Disclosures: Pratt: CSL Behring: Research Funding; Bayer Healthcare: Research Funding; Baxter: Honoraria; Grifols: Honoraria.

scholarly journals Detection and Molecular Characterization of Benzimidazole Resistance Among Colletotrichum truncatum Isolates Infecting Bell Pepper in Trinidad

Plant Disease ◽  
2016 ◽  
Vol 100 (6) ◽  
pp. 1146-1152 ◽  
Author(s):  
H. Ramdial ◽  
F. N. Hosein ◽  
S. N. Rampersad
Keyword(s):  
Amino Acid ◽  
Colony Growth ◽  
Bell Pepper ◽  
Single Amino Acid ◽  
Bootstrap Support ◽  
Amino Acid Substitutions ◽  
Nucleotide Polymorphisms ◽  
Colletotrichum Truncatum ◽  
Accurate Identification ◽  
Β Tubulin

Anthracnose is an economically important disease that affects pepper (Capsicum spp.) production worldwide. Eighty-seven Colletotrichum truncatum isolates infecting bell pepper in Trinidad were isolated and screened for resistance to benomyl. All isolates were found to be highly resistant at the discriminatory dose of 10.0 μg/ml. The effective concentration required to achieve 50% colony growth inhibition values were found to be significantly higher (P ≤ 0.05) for isolates collected in South Trinidad compared with those collected in North Trinidad. Isolates with the resistant phenotype had a single amino acid substitution from glutamic acid to alanine at position 198 (E198A) within the β-tubulin 2 gene. Single-nucleotide polymorphisms that result in amino acid substitutions in the β-tubulin 2 protein are associated with high resistance to benzimidazole chemistries. There were also two other deduced amino acid changes at nucleotide positions 359 to 361 (ATA/TTG [F270Y]) and at 362 to 364 (CGC/GCC [A271S]). Genetic analysis revealed that benomyl-sensitive isolates clustered separately from the resistant isolates regardless of species, with strong bootstrap support (85%). Within the resistance cluster, there was an apparent differentiation among those isolates with the F200Y, E198A, and E198K genotypes, with moderate support (>60%) for clustering of the F200Y and E198K genotypes. C. truncatum clustered separately (97%) from the other resistant genotypes due to the additional amino acid substitutions detected. The findings also indicated that consistent benzimidazole fungicide use may explain the predominance of the C. truncatum pathogen populations in bell pepper fields in Trinidad because sensitive C. gloeosporioides sensu lato isolates were selectively minimized. This underlines the importance of accurate identification of Colletotrichum spp. associated with anthracnose disease and routine monitoring for development of fungicide resistance.

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