Glucose Modulation of the Local Conformational Flexibility and Free-Energy Basin Shape of Human and Yeast Hexokinases: A Comparative Molecular Dynamics Analysis

Human hexokinases HK1 (H) and HK2 (H) are cancer drug targets, yet yeast orthologs often serve as computational surrogates. We analyzed three independent 100 ns molecular dynamics replicates of HK1 (H), HK2 (H) and their Saccharomyces cerevisiae homologs [HK1 (Y), HK2 (Y)] in free and glucose-bound states. Because this window samples the local free-energy basin rather than the microsecond-scale open↔closed transition, we compared how glucose modulates local flexibility and basin shape, combining potentials of mean force along the radius of gyration and backbone RMSD with basin occupancy, residence times, local well stiffness (kloc) and volumetric compliance. In both human enzymes glucose tended to enlarge volumetric fluctuations (compliance 1.8–3.2-fold higher); HK1 (H) compacted progressively, to a replicate-dependent extent, whereas HK2 (H) showed the most reproducible effect: lower free-state basin occupancy, a 2.8-fold broader RMSD distribution and a threefold softer well (kloc 980 ± 145 versus 295 ± 112 kBT/nm2). The yeast enzymes responded weakly or oppositely: in HK1 (Y) glucose lowered backbone deviation and narrowed within-replicate fluctuations (compliance 2.5-fold lower), whereas HK2 (Y) changed minimally. In these simulations, the glucose–flexibility coupling was not conserved between isoforms or species, supporting human-specific structural ensembles, rather than yeast models, for computational inhibitor design.