Beyond MIDAS: An In Silico Study of a Putative Noncanonical C16 Binding Site in αvβ3 Integrin

Integrins are essential transmembrane receptors that mediate bidirectional signaling between the extracellular matrix (ECM) and the cytoskeleton. Among them, αvβ3 and α5β1 are key regulators of angiogenesis, metastasis, and tumor progression. The laminin-1-derived C16 peptide (KAFDITYVRLKF) interacts with both integrins, but the molecular determinants governing its specificity remain unclear. Here, we applied an integrative computational approach combining ADMET predictions, molecular docking, and all-atom molecular dynamics (MD) simulations to characterize C16 pharmacokinetics and its predicted binding modes to αvβ3 and α5β1. In silico pharmacological profiling indicated that C16 displays low oral bioavailability and limited permeability but possesses favorable binding properties and moderate metabolic stability. Docking and MD analyses revealed that C16 interacts with the canonical metal ion-dependent adhesion site (MIDAS) in both integrins via conserved hydrogen bonds and hydrophobic contacts. Strikingly, an additional putative noncanonical binding pocket (S2) was identified in αvβ3 (not in α5β1), located opposite the MIDAS. This site forms an amphiphilic interface stabilized by complementary electrostatic and hydrophobic interactions, providing a structural rationale for the preferential interaction of C16 with αvβ3. MD simulations confirmed the stability of both S1 and S2 complexes, with reduced conformational fluctuations and lower binding free energies for αvβ3, particularly at the S2 site. These computational findings propose a novel αvβ3 C16 recognition mode, offering computational insights into predicted molecular determinants of integrin subtype selectivity and informing the rational design of peptide-based therapeutics targeting angiogenesis and tumor invasion.