Structure, Chemical Reactivity, and In Silico Noncovalent PBP2a Allosteric Binding of an Isoxazolyl‐Benzodiazepinone Hybrid: A Crystallographic and Computational Study

ABSTRACT

Methicillin‐resistant Staphylococcus aureus (MRSA) remains a major clinical challenge due to β ‐lactam resistance mediated by penicillin‐binding protein 2a (PBP2a), prompting growing interest in allosteric modulation as an alternative therapeutic strategy. In this study, the structure, reactivity, and prospective PBP2a allosteric binding potential of the heterocyclic hybrid 4‐(2‐((3‐(4‐chlorophenyl) isoxazol‐5‐yl)methoxy)phenyl)‐1‐methyl‐1,5‐benzodiazepin‐2‐one are investigated through combined crystallographic and computational methods. The solid‐state molecular conformation and crystal structure, determined by single‐crystal x‐ray diffraction, reveal a stabilized supramolecular architecture sustained by nonclassical C─H⋯O and C─H⋯Cl hydrogen bonds, C─H⋯ π contacts, and π – π stacking interactions. The nature and energetics of these interactions are further examined using density functional theory (DFT)‐based quantum theory of atoms in molecule (QTAIM) and independent gradient model (IGM) analyses. Furthermore, conceptual DFT descriptors characterize the molecule as a strong electrophile with moderate‐to‐strong nucleophilic character, while Parr functions and molecular electrostatic potential (MEP) maps identify site‐specific reactivity. Molecular docking shows stable noncovalent binding within the PBP2a allosteric site, with a binding affinity of −7.4 kcal/mol, involving key N─H⋯O hydrogen bonds and T‐shaped π – π stacking with Lys273 and Tyr297, comparable to those observed for the reference modulator ceftaroline ( CFT ). Complementary Allosteric Signaling and Mutation Analysis (AlloSigMA) further predicts long‐range energetic coupling between the BZD‐binding pocket and the catalytic region of PBP2a.