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The Gln233Arg (A>G; rs1137101) polymorphism of the leptin receptor gene (LEPR) has been investigated extensively and is reported to be associated with different metabolic states. In this investigation, we aimed to study the frequency of Gln233Arg genotypes and alleles in a group of Saudi women stratified by their body mass index (BMI), to correlate the LEPR genotypes with variations in anthropometric, lipid and hormonal parameters and to investigate conformational and structural variations in the mutant LEPR using molecular dynamic (MD) investigations. The study group included 122 Saudi women (normal weight = 60; obese = 62) attending the clinics for a routine checkup. Anthropometric data: height, weight, waist and hip circumference were recorded and fasting serum sample was used to estimate glucose, lipids, ghrelin, leptin and insulin. BMI, W/H ratio, and HOMA-IR values were calculated. Whole blood sample was used to extract DNA; exon 6 of the LEPR gene was amplified by PCR and sequencing was conducted on an ABI 3100 Avant Genetic Analyser. Molecular Dynamic Simulation studies were carried out using different softwares. The results showed the presence of all three genotypes of Gln233Arg in Saudi women, but the frequencies were significantly different when compared to reports from some populations. No differences were seen in the genotype and allele frequencies between the normal weight and obese women. Stratification by the genotypes showed significantly higher BMI, waist and hip circumference, leptin, insulin, fasting glucose and HOMA-IR and lower ghrelin levels in obese women carrying the GG genotype. Even in the normal weight group, individuals with GG genotype had higher BMI, waist and hip circumference and significantly lower ghrelin levels. The MD studies showed a significant effect of the Gln/Arg substitution on the conformation, flexibility, root-mean-square fluctuation (RMSF), radius of gyration (Rg) values, solvent-accessible surface area (SASA) and number of inter- and intra-molecular H-bonds. The results suggest that the structural changes brought about by the mutation, influence the signaling pathways by some unknown mechanism, which may be contributing to the abnormalities seen in the individuals carrying the G allele of rs1137101.

Computational prediction of cancer associated SNPs from the large pool of SNP dataset is now being used as a tool for detecting the probable oncogenes, which are further examined in the wet lab experiments. The lack in prediction accuracy has been a major hurdle in relying on the computational results obtained by implementing multiple tools, platforms and algorithms for cancer associated SNP prediction. Our result obtained from the initial computational compilations suggests the strong chance of Aurora-A G325W mutation (rs11539196) to cause hepatocellular carcinoma. The implementation of molecular dynamics simulation (MDS) approaches has significantly aided in raising the prediction accuracy of these results, but measuring the difference in the convergence time of mutant protein structures has been a challenging task while setting the simulation timescale. The convergence time of most of the protein structures may vary from 10 ns to 100 ns or more, depending upon its size. Thus, in this work we have implemented 200 ns of MDS to aid the final results obtained from computational SNP prediction technique. The MDS results have significantly explained the atomic alteration related with the mutant protein and are useful in elaborating the change in structural conformations coupled with the computationally predicted cancer associated mutation. With further advancements in the computational techniques, it will become much easier to predict such mutations with higher accuracy level.

Ras-related C3 botulinum toxin substrate 1 (RAC1) is a plasma membrane-associated small GTPase which cycles between the active GTP-bound and inactive GDP-bound states. There is wide range of evidences indicating its active participation in inducing cancer-associated phenotypes. RAC1 F28L mutation (RAC(F28L)) is a fast recycling mutation which has been implicated in several cancer associated cases. In this work we have performed molecular docking and molecular dynamics simulation (~0.3 μs) to investigate the conformational changes occurring in the mutant protein. The RMSD, RMSF and NHbonds results strongly suggested that the loss of native conformation in the Switch I region in RAC1 mutant protein could be the reason behind its oncogenic transformation. The overall results suggested that the mutant protein attained compact conformation as compared to the native. The major impact of mutation was observed in the Switch I region which might be the crucial reason behind the loss of interaction between the guanine ring and F28 residue.

BACKGROUND: Aurora kinases belong to the highly conserved kinase family and play a vital role in cell cycle regulation. The structure and function of these kinases are inter-related and sometimes they also act as substitutes in case of knockdown of other aurora kinases. METHOD: In this work we carried out the evolutionary reconstruction and population genetic studies of aurora kinase proteins. Substitution saturation test, CAI (Codon adaptation index), gene expression and RSCU (Relative synonymous codon usage) values were computed for all the three aurora kinases. Linear regression method was used to check the dependency of gene expression on their CAI values. RESULTS: The results suggested that aurora-B and aurora-C has shown convergence in their evolutionary pathway. Moreover, the aurora-A I57V mutation showed high penetrance in human population and exist at very high frequency (84.4%) when compared to the native residue (15.6%). The mutation showed notable range of functional gain and seemed to be promising for the evolution of aurora-A function. Mutant allele might also become a challenging prospect for understanding the pattern of evolution followed by cell cycle kinases. CONCLUSION: The overall result suggested that the aurora-A is currently under the evolutionary transition and to determine the functional significance of the mutation further investigation are required.

BACKGROUND: AKT1 (v-akt murine thymoma viral oncogene homologue 1) kinase is one of the most frequently activated proliferated and survival pathway of cancer. Recently it has been shown that E17K mutation in the Pleckstrin Homology (PH) domain of AKT1 protein leads to cancer by amplifying the phosphorylation and membrane localization of protein. The mutant has shown resistance to AKT1/2 inhibitor VIII drug molecule. In this study we have demonstrated the detailed structural and molecular consequences associated with the activity regulation of mutant protein. METHODS: The docking score exhibited significant loss in the interaction affinity to AKT1/2 inhibitor VIII drug molecule. Furthermore, the molecular dynamics simulation studies presented an evidence of rapid conformational drift observed in mutant structure. RESULTS: There was no stability loss in mutant as compared to native structure and the major cation-π interactions were also shown to be retained. Moreover, the active residues involved in membrane localization of protein exhibited significant rise in NHbonds formation in mutant. The rise in NHbond formation in active residues accounts for the 4-fold increase in the membrane localization potential of protein. CONCLUSION: The overall result suggested that, although the mutation did not induce any stability loss in structure, the associated pathological consequences might have occurred due to the rapid conformational drifts observed in the mutant AKT1 PH domain. GENERAL SIGNIFICANCE: The methodology implemented and the results obtained in this work will facilitate in determining the core molecular mechanisms of cancer-associated mutations and in designing their potential drug inhibitors.