Harnessing virtual screening and MD simulations: a multistage approach to identifying potent and nontoxic agonists...

Obesity-induced insulin resistance impairs glucose tolerance and β-cell function, significantly contributing to the pathogenesis of type 2 diabetes (T2D). Protein kinase A (PKA), being one of the key effector molecules of the cyclic AMP (cAMP) pathway, increases insulin secretion via membrane activity, gene expression, and exocytosis of insulin granules. The previous studies were limited to either target cAMP analogs as PKA agonist or mostly flavonoids using In vivo and In vitro studies (Hameed in Int J Biol Macromol 119:149-156, 2018;Shahab in Biomed Pharmacother 177, 2024;Hameed in Eur J Pharmacol 820:245-255, 2018;Hameed in Eur J Pharmacol 858, 2019;Hafizur in Med Chem Res 27:1408-1418, 2018;). To speed up the process, this study aimed to identify potential PKA activators as therapeutic agents for restoring β-cell function in Type 2 Diabetes (T2D) using a multistage virtual screening approach. In the initial phase, a ligand-based pharmacophore model was constructed to screen an in-house small molecule database for potential PKA agonists. By targeting the essential pharmacophoric features necessary for interaction with the cyclic nucleotide-binding (CNB) domain of PKA, the goal was to identify compounds with strong binding affinities and therapeutic promise. To gain deeper insights into the molecular mechanisms of PKA activation and evaluate key interactions and dynamic stability, a subset of promising hits was subjected to all-atom molecular dynamics simulations. Simulations showed significant conformational changes in PKA complexes, with average backbone root mean square deviations (RMSD) of 0.37 ± 0.15 nm for Comp-03, 0.53 ± 0.18 nm for Comp-11, 0.31 ± 0.06 nm for Comp-17, 0.28 ± 0.03 nm for Comp-38, and 0.48 ± 0.13 nm for Comp-41. The N3A motif showed consistent fluctuations, suggesting increased flexibility. Binding free energy calculations showed binding free energies (ΔGbind) for cAMP, Comp-03, Comp-17, Comp-38, and Comp-41, with ΔGbind values of - 62.87 ± 10.04, - 68.57 ± 12.77, - 78.13 ± 16.36, - 62.67 ± 13.06, and - 80.87 ± 10.45 kcal/mol, respectively. To further probe the conformational stability of these complexes, multidimensional scaling and free energy profiling were carried out. This exhaustive research study, involving examination of stability dynamics, deviation patterns, interaction networks, conformational changes, and energy profiles, provides profound understanding about mechanisms that activate PKA. The findings highlight several promising lead compounds, notably Comp-03, Comp-17, Comp-38, and Comp-41, which exhibit superior potential to activate PKA compared to cAMP. These findings lay a strong foundation for the development of novel PKA activators as potential therapeutic agents for managing T2D.