Computational analysis of non-synonymous SNPs in the sheep MC4R gene

The melanocortin-4 receptor (MC4R) gene is central to appetite regulation, body weight, and energy balance in vertebrates, with significant influence on sheep growth, fat deposition, feed efficiency, and reproduction. However, the functional consequences of non-synonymous single nucleotide polymorphisms (nsSNPs) in this gene remain unclear. This study aimed to evaluate the structural and functional impact of non-synonymous single nucleotide polymorphisms (nsSNPs) in the ovine MC4R gene using an integrated computational approach. A set of nsSNPs was analyzed to identify potentially deleterious variants based on their predicted effects on protein function, stability, and evolutionary conservation. A total of 12 ovine MC4R nsSNPs were identified from Ensembl and UniPro , analyzed using an integrated computational framework.. Functional effects were predicted by SIFT, PolyPhen-2, SNPs&GO, and PhD-SNP, while protein stability was assessed with I-Mutant, MUpro, CUPSAT, and DynaMut. Conservation analysis (ConSurf), structural modeling (SOPMA, I-TASSER, GalaxyWEB), validation (PROCHECK, ProSA), and molecular dynamics simulations (100 ns, WEBGRO) were performed. Four nsSNPs (R220G, G98R, E100K, Y187C) were consistently predicted as deleterious, and damaging nsSNPs often occurred in conserved regions. . Structural modeling and molecular dynamics simulations revealed that these nsSNPs may alter protein stability, flexibility, and conformational dynamics compared to the wild-type protein.. These findings identify several nsSNPs as potential candidate variants that may influence MC4R structure and function. However, their biological and practical relevance should be interpreted with caution, as the results are based solely on computational predictions. The identified variants represent promising targets for further investigation, but their application in selective breeding programs requires validation through population-based studies, genotype-phenotype association analyses, and experimental approaches. Overall, this study provides a framework for prioritizing potentially functional MC4R variants, rather than direct evidence for their use in breeding applications.