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Investigation on the role of nsSNPs in HNPCC genes--a bioinformatics approach.

Doss CG, Sethumadhavan R - J. Biomed. Sci. (2009)

Bottom Line: Missense mutations are nucleotide substitutions that change an amino acid in a protein, the deleterious effects of these mutations are commonly attributed to their impact on primary amino acid sequence and protein structure.The PupaSuite predicted the phenotypic effect of SNPs on the structure and function of the affected protein.Our study demonstrates the presence of other deleterious mutations and also endorses with in vivo experimental studies.

View Article: PubMed Central - HTML - PubMed

Affiliation: Bioinformatics Division, School of Biotechnology, Chemical and Biomedical Engineering, Vellore Institute of Technology, Vellore 632014, Tamil Nadu, India. georgecp77@yahoo.co.in

ABSTRACT

Background: A central focus of cancer genetics is the study of mutations that are causally implicated in tumorigenesis. The identification of such causal mutations not only provides insight into cancer biology but also presents anticancer therapeutic targets and diagnostic markers. Missense mutations are nucleotide substitutions that change an amino acid in a protein, the deleterious effects of these mutations are commonly attributed to their impact on primary amino acid sequence and protein structure.

Methods: The method to identify functional SNPs from a pool, containing both functional and neutral SNPs is challenging by experimental protocols. To explore possible relationships between genetic mutation and phenotypic variation, we employed different bioinformatics algorithms like Sorting Intolerant from Tolerant (SIFT), Polymorphism Phenotyping (PolyPhen), and PupaSuite to predict the impact of these amino acid substitutions on protein activity of mismatch repair (MMR) genes causing hereditary nonpolyposis colorectal cancer (HNPCC).

Results: SIFT classified 22 of 125 variants (18%) as 'Intolerant." PolyPhen classified 40 of 125 amino acid substitutions (32%) as "Probably or possibly damaging". The PupaSuite predicted the phenotypic effect of SNPs on the structure and function of the affected protein. Based on the PolyPhen scores and availability of three-dimensional structures, structure analysis was carried out with the major mutations that occurred in the native protein coded by MSH2 and MSH6 genes. The amino acid residues in the native and mutant model protein were further analyzed for solvent accessibility and secondary structure to check the stability of the proteins.

Conclusion: Based on this approach, we have shown that four nsSNPs, which were predicted to have functional consequences (MSH2-Y43C, MSH6-Y538S, MSH6-S580L, and MSH6-K854M), were already found to be associated with cancer risk. Our study demonstrates the presence of other deleterious mutations and also endorses with in vivo experimental studies.

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(A) Native structure of MSH6 gene with 'B' chain of PDB ID 2O8C(orange). (B) Superimposed structure of native tyrosine (orange) with mutant amino acid serine (pale green) at 538 position in 2O8C with RMSD 3.52 Å. (C) Superimposed structure of native serine (orange) with mutant amino acid leucine (pale green) at 580 position in 2O8C with RMSD 3.37 Å. (D)Superimposed structure of native lysine (orange) with mutant amino acid methionine (pale green) at 854 position in 2O8C with RMSD 3.30 Å.
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Figure 3: (A) Native structure of MSH6 gene with 'B' chain of PDB ID 2O8C(orange). (B) Superimposed structure of native tyrosine (orange) with mutant amino acid serine (pale green) at 538 position in 2O8C with RMSD 3.52 Å. (C) Superimposed structure of native serine (orange) with mutant amino acid leucine (pale green) at 580 position in 2O8C with RMSD 3.37 Å. (D)Superimposed structure of native lysine (orange) with mutant amino acid methionine (pale green) at 854 position in 2O8C with RMSD 3.30 Å.

Mentions: Single amino acid mutations can significantly change the stability of a protein structure. So, the knowledge of a protein's three-dimensional (3D) structure is essential for a full understanding of its functionality. Mapping the deleterious nsSNPs into protein structure information was obtained from dbSNP and SAAPdb. The available structure for the MSH2 and MSH6 gene is reported to have a PDB ID (2O8C). Mutation analysis was performed based on the results obtained from highest PolyPhen scores. The mutations for 2O8C at their corresponding positions were performed by SWISS-PDB viewer independently to achieve modeled structures. Then, energy minimizations were performed by NOMAD-Ref server for the native type protein 2O8C and the mutant type structures. It can be inferred from (Table 1) that nsSNPs in MSH2 gene with ids namely rs17217723, rs180522 and rs41294982 showed the highest PolyPhen scores 2.970, 3.352 and 3.379 respectively. According to this, the mutation occurred for native protein in the 'A' chain of PDB ID 2O8C at position Y43C with an SNP ID (rs17217723), H639Q with an SNP ID (rs180522) and P670L with an SNP ID (rs41294982) based on PolyPhen results. It can be seen that the total energy for mutant type structure Y43C, H639Q and P670L were found to be -53305.15, -53377.01, -53405.59 Kcal/mol respectively. The RMSD values between the native type (2O8C) and the mutant Y43C is 4.30 Å, between native type and the mutant H639Q is 3.93 Å and between native type and the mutant P670L is 3.65 Å. The total energy and RMSD value of mutant structure Y43C is high when compared to the other mutants H639Q and P670L respectively. Similarly, for MSH6 gene based on the PolyPhen scores, mutation analysis was performed in nsSNPs with IDs namely rs728619, rs41295270 and rs34374438 respectively. According to this, the mutation occurred for native protein in the 'B' chain of PDB ID 2O8C at position Y538S with an SNP ID (rs728619), S580L with an SNP ID (rs41295270) and K854M with an SNP ID (rs34374438). It can be seen that the total energy for mutant type structure Y538S, S580L and K854M were found to be -58509.39, -58513.55, -58506.94 Kcal/mol respectively. The RMSD values between the native type (2O8C) and the mutant Y538S is 3.52 Å, between native type and the mutant S580L is 3.37 Å and between native type and the mutant K854M is 3.30 Å. The total energy and RMSD value of mutant structure Y43C is high when compared to the other mutants H639Q and P670L in MSH2 gene, while all the three mutants Y538S, S580L and K854M in MSH6 showed almost same total energy and RMSD. Higher the RMSD value more will be the deviation between native and mutant type structures and which in turn changes their functional activity. The superimposed structures of the native protein 2O8C (chain A) with the three mutant type proteins Y43C, H639Q and P670L of MSH2 gene are shown in shown in (Figure 2a, b, c &2d) and the superimposed structures of the native protein 2O8C (chain B) with the three mutant type proteins Y538S, S580L and K854M of MSH6 gene are shown in (Figure 3a, b, c &3d) respectively.


Investigation on the role of nsSNPs in HNPCC genes--a bioinformatics approach.

Doss CG, Sethumadhavan R - J. Biomed. Sci. (2009)

(A) Native structure of MSH6 gene with 'B' chain of PDB ID 2O8C(orange). (B) Superimposed structure of native tyrosine (orange) with mutant amino acid serine (pale green) at 538 position in 2O8C with RMSD 3.52 Å. (C) Superimposed structure of native serine (orange) with mutant amino acid leucine (pale green) at 580 position in 2O8C with RMSD 3.37 Å. (D)Superimposed structure of native lysine (orange) with mutant amino acid methionine (pale green) at 854 position in 2O8C with RMSD 3.30 Å.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2682794&req=5

Figure 3: (A) Native structure of MSH6 gene with 'B' chain of PDB ID 2O8C(orange). (B) Superimposed structure of native tyrosine (orange) with mutant amino acid serine (pale green) at 538 position in 2O8C with RMSD 3.52 Å. (C) Superimposed structure of native serine (orange) with mutant amino acid leucine (pale green) at 580 position in 2O8C with RMSD 3.37 Å. (D)Superimposed structure of native lysine (orange) with mutant amino acid methionine (pale green) at 854 position in 2O8C with RMSD 3.30 Å.
Mentions: Single amino acid mutations can significantly change the stability of a protein structure. So, the knowledge of a protein's three-dimensional (3D) structure is essential for a full understanding of its functionality. Mapping the deleterious nsSNPs into protein structure information was obtained from dbSNP and SAAPdb. The available structure for the MSH2 and MSH6 gene is reported to have a PDB ID (2O8C). Mutation analysis was performed based on the results obtained from highest PolyPhen scores. The mutations for 2O8C at their corresponding positions were performed by SWISS-PDB viewer independently to achieve modeled structures. Then, energy minimizations were performed by NOMAD-Ref server for the native type protein 2O8C and the mutant type structures. It can be inferred from (Table 1) that nsSNPs in MSH2 gene with ids namely rs17217723, rs180522 and rs41294982 showed the highest PolyPhen scores 2.970, 3.352 and 3.379 respectively. According to this, the mutation occurred for native protein in the 'A' chain of PDB ID 2O8C at position Y43C with an SNP ID (rs17217723), H639Q with an SNP ID (rs180522) and P670L with an SNP ID (rs41294982) based on PolyPhen results. It can be seen that the total energy for mutant type structure Y43C, H639Q and P670L were found to be -53305.15, -53377.01, -53405.59 Kcal/mol respectively. The RMSD values between the native type (2O8C) and the mutant Y43C is 4.30 Å, between native type and the mutant H639Q is 3.93 Å and between native type and the mutant P670L is 3.65 Å. The total energy and RMSD value of mutant structure Y43C is high when compared to the other mutants H639Q and P670L respectively. Similarly, for MSH6 gene based on the PolyPhen scores, mutation analysis was performed in nsSNPs with IDs namely rs728619, rs41295270 and rs34374438 respectively. According to this, the mutation occurred for native protein in the 'B' chain of PDB ID 2O8C at position Y538S with an SNP ID (rs728619), S580L with an SNP ID (rs41295270) and K854M with an SNP ID (rs34374438). It can be seen that the total energy for mutant type structure Y538S, S580L and K854M were found to be -58509.39, -58513.55, -58506.94 Kcal/mol respectively. The RMSD values between the native type (2O8C) and the mutant Y538S is 3.52 Å, between native type and the mutant S580L is 3.37 Å and between native type and the mutant K854M is 3.30 Å. The total energy and RMSD value of mutant structure Y43C is high when compared to the other mutants H639Q and P670L in MSH2 gene, while all the three mutants Y538S, S580L and K854M in MSH6 showed almost same total energy and RMSD. Higher the RMSD value more will be the deviation between native and mutant type structures and which in turn changes their functional activity. The superimposed structures of the native protein 2O8C (chain A) with the three mutant type proteins Y43C, H639Q and P670L of MSH2 gene are shown in shown in (Figure 2a, b, c &2d) and the superimposed structures of the native protein 2O8C (chain B) with the three mutant type proteins Y538S, S580L and K854M of MSH6 gene are shown in (Figure 3a, b, c &3d) respectively.

Bottom Line: Missense mutations are nucleotide substitutions that change an amino acid in a protein, the deleterious effects of these mutations are commonly attributed to their impact on primary amino acid sequence and protein structure.The PupaSuite predicted the phenotypic effect of SNPs on the structure and function of the affected protein.Our study demonstrates the presence of other deleterious mutations and also endorses with in vivo experimental studies.

View Article: PubMed Central - HTML - PubMed

Affiliation: Bioinformatics Division, School of Biotechnology, Chemical and Biomedical Engineering, Vellore Institute of Technology, Vellore 632014, Tamil Nadu, India. georgecp77@yahoo.co.in

ABSTRACT

Background: A central focus of cancer genetics is the study of mutations that are causally implicated in tumorigenesis. The identification of such causal mutations not only provides insight into cancer biology but also presents anticancer therapeutic targets and diagnostic markers. Missense mutations are nucleotide substitutions that change an amino acid in a protein, the deleterious effects of these mutations are commonly attributed to their impact on primary amino acid sequence and protein structure.

Methods: The method to identify functional SNPs from a pool, containing both functional and neutral SNPs is challenging by experimental protocols. To explore possible relationships between genetic mutation and phenotypic variation, we employed different bioinformatics algorithms like Sorting Intolerant from Tolerant (SIFT), Polymorphism Phenotyping (PolyPhen), and PupaSuite to predict the impact of these amino acid substitutions on protein activity of mismatch repair (MMR) genes causing hereditary nonpolyposis colorectal cancer (HNPCC).

Results: SIFT classified 22 of 125 variants (18%) as 'Intolerant." PolyPhen classified 40 of 125 amino acid substitutions (32%) as "Probably or possibly damaging". The PupaSuite predicted the phenotypic effect of SNPs on the structure and function of the affected protein. Based on the PolyPhen scores and availability of three-dimensional structures, structure analysis was carried out with the major mutations that occurred in the native protein coded by MSH2 and MSH6 genes. The amino acid residues in the native and mutant model protein were further analyzed for solvent accessibility and secondary structure to check the stability of the proteins.

Conclusion: Based on this approach, we have shown that four nsSNPs, which were predicted to have functional consequences (MSH2-Y43C, MSH6-Y538S, MSH6-S580L, and MSH6-K854M), were already found to be associated with cancer risk. Our study demonstrates the presence of other deleterious mutations and also endorses with in vivo experimental studies.

Show MeSH
Related in: MedlinePlus