Probing the Molecular Interactions of A22 with Prokaryotic Actin MreB and Eukaryotic Actin: A Computational and Experimental Study.

Actin is a major cytoskeletal system that mediates the intricate organization of macromolecules within cells. The bacterial cytoskeletal protein MreB is a prokaryotic actin-like protein governing the cell shape and intracellular organization in many rod-shaped bacteria, including pathogens. MreB stands as a target for antibiotic development, and compounds like A22 and its analogue, MP265, are identified as potent inhibitors of MreB. The bacterial actin MreB shares structural homology with eukaryotic actin despite lacking sequence similarity. It is currently not clear whether small molecules that inhibit MreB can act on eukaryotic actin due to their structural similarity. In this study, we investigate the molecular interactions between A22 and its analogue MP265 with MreB and eukaryotic actin through a molecular dynamics approach. Employing MD simulations and free energy calculations with an all-atom model, we unveil the robust interaction of A22 and MP265 with MreB, and substantial binding affinity is observed for A22 and MP265 with eukaryotic actin. Experimental assays reveal A22's toxicity to eukaryotic cells, including yeast and human glioblastoma cells. Microscopy analysis demonstrates the profound effects of A22 on actin organization in human glioblastoma cells. This integrative computational and experimental study provides new insights into A22's mode of action, highlighting its potential as a versatile tool for probing the dynamics of both prokaryotic and eukaryotic actins.