Comprehensive analysis of non-synonymous single-nucleotide polymorphism of human TSC1 and TSC2 genes: An in silico...

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder resulting from the mutations in the TSC1 and TSC2 genes and is distinguished by benign hamartoma formation in multiple organs. The TSC1-TSC2 complex regulates mTORC1 signaling in response to cellular growth conditions. This study aims to predict the structural stability and functional effects of non-synonymous single-nucleotide polymorphisms (nsSNPs) in human TSC1 and TSC2 using computational approaches. Twelve computational tools were assessed using receiver operating characteristic (ROC) analysis and applied to identify deleterious nsSNPs. Protein stability was predicted using I-Mutant 2.0 and MUpro, while evolutionary conservation was analyzed with ConSurf. NetPhos 3.1 identified potential PTM sites, and MutPred2.0 evaluated their functional impact. Project HOPE assessed mutation-induced physicochemical changes. Structural models were validated using multiple tools, visualized in ChimeraX 1.9, and further evaluated by molecular dynamics simulation to confirm wild-type and mutant stability. A combined in silico analysis identified twelve high-risk nsSNPs in TSC1 and sixteen in TSC2, all reducing protein stability, located in conserved regions, and potentially disrupting phosphorylation sites. MutPred and Project HOPE confirmed their impact on protein function. Functional analysis showed TSC1 and TSC2 affect mTORC1 and PI3K-Akt pathways. RMSF and RMSD analyses revealed that TSC1 variants rs1846545280 (G236E), and rs2132135678 (V234E), and TSC2 variants rs45517223 (S758C), rs2151354925 (T836P), and rs45517365 (R1570W) had the largest structural fluctuations. Substitution with glutamic acid, a negatively charged and bulkier residue, may disrupt local folding of TSC1. Similarly, replacement of arginine with tyrosine at position 1570 may impair Rheb binding at the GAP domain of TSC2. These findings highlight potentially pathogenic nsSNPs in TSC1 and TSC2.