Date of Award
Spring 2024
Document Type
Thesis
Degree Name
Master of Science (MS) in Biology
Department
Biology
Committee Chair
Dr. Ashley Turner
Abstract
Type 2 diabetes mellitus (T2DM) is hallmarked by insulin resistance, with the INSR gene identified as a key player in this condition in humans. This gene is known to harbor genetic variants with a wide range of clinical significance from pathogenic to variants of uncertain significance (VUS) to benign. This project investigates a VUS associated with T2DM identified through ClinVar. A gene mutational analysis, predictive amino acid substitution analysis, and protein modeling predict INSR c.1628C>T (p. Thr543Met) to be likely pathogenic or damaging. PolyPhen-2 predicts this variant to be probably damaging (HumDiv score of 1.000).
Evolutionary conservation of the VUS loci across multiple species was confirmed through multiple sequence alignment carried out through Benchling. The VUS results in an amino acid class change from the polar, hydrophilic amino acid threonine to the nonpolar, hydrophobic amino acid methionine. Utilizing SWISS-MODEL, homology-based protein models were generated for both human native and VUS mutant INSR proteins. Subsequent analysis in PyMOL revealed an RMSD value of 0.001. The affected amino acid resides in the extracellular alpha subunit, which may contribute to its potential impact to protein structure and function. To understand the functional implications of this structural alteration, we plan to observe potential phenotypic differences between wildtype N2 C. elegans nematodes and a CRISPR-Cas9 engineered daf-2 VUS nematodes. This research aims to elucidate the functional impact of the INSR variant and provide valuable insight into its role in the context of insulin resistance and T2DM.