Characterizing the Molecular Mechanism of the Lethal C423D Mutation in FgMyoI: A Molecular Perspective.

The lethal mutation C423D in Fusarium graminearum myosin I (FgMyoI) occurs close to the binding pocket of the allosteric inhibitor phenamacril and causes severe inhibition on mycelial growth of F. graminearum strain PH-1. Here, based on extensive Gaussian accelerated molecular dynamics simulations and wet experiments, we elucidate the underlying molecular mechanism of the abnormal functioning of the FgMyoIC423D mutant at the atomistic level. Our results suggest that the damaging mutation C423D exhibits a synergistic allosteric inhibition mechanism similar to but more robust than that of phenamacril, including effects on the active site and actin binding. Unlike phenamacril-induced closure of Switch2, the mutation results in unfolding of the N-terminal relay helix with a partially opened Switch2 and blocks the structural rearrangement of the relay/SH1 helices, impairing the proper initiation of the recovery stroke. Due to the significant influence of C423D mutation on the function of FgMyoI, designing covalent inhibitors targeting this site holds tremendous potential.

Allosteric inhibition of myosin by phenamacril: a synergistic mechanism revealed by computational and experimental approaches.

BACKGROUND: Myosin plays a crucial role in cellular processes, while its dysfunction can lead to organismal malfunction. Phenamacril (PHA), a highly species-specific and non-competitive inhibitor of myosin I (FgMyoI) from Fusarium graminearum, has been identified as an effective fungicide for controlling plant diseases caused by partial Fusarium pathogens, such as wheat scab and rice bakanae. However, the molecular basis of its action is still unclear. RESULTS: This study used multiple computational approaches first to elucidate the allosteric inhibition mechanism of FgMyoI by PHA at the atomistic level. The results indicated the increase of adenosine triphosphate (ATP) binding affinity upon PHA binding, which might impede the release of hydrolysis products. Furthermore, simulations revealed a broadened outer cleft and a significantly more flexible interface for actin binding, accompanied by a decrease in signaling transduction from the catalytic center to the actin-binding interface. These various effects might work together to disrupt the actomyosin cycle and hinder the ability of motor to generate force. Our experimental results further confirmed that PHA reduces the enzymatic activity of myosin and its binding with actin. CONCLUSION: Therefore, our findings demonstrated that PHA might suppress the function of myosin through a synergistic mechanism, providing new insights into myosin allostery and offering new avenues for drug/fungicide discovery targeting myosin. © 2023 Society of Chemical Industry.