Molecular dynamics simulations integrating kinetics for Pb2+-induced arginine kinase inactivation and aggregation

Abstract We built a 3D structure of arginine kinase from Exopalaemon carinicauda (ECAK) on the basis of the ORF gene sequence to conduct molecular dynamics simulations between ECAK and Pb 2+ . As a result, the binding mechanism of Pb 2+ to ECAK was predicted from the binding sites, and the structural change was elucidated. To confirm the simulated results, the Pb 2+ -mediated inhibition and aggregation of ECAK were subsequently conducted. We found that Pb 2+ partially inactivated the activity of ECAK with relatively strong binding. The spectrofluorimetry results showed that Pb 2+ induced tertiary structural changes of ECAK, with the substantial exposure of hydrophobic surfaces directly induced by ECAK aggregation. The ECAK aggregation process induced by Pb 2+ occurred with multi-phase kinetics. The addition of osmolytes did not protect ECAK from Pb 2+ inactivation. Because AK plays an important role in the cellular energy metabolism of invertebrates, our study suggests new information about the effect of Pb 2+ on ECAK's enzymatic function and unfolding, including aggregation, which may be toxic to invertebrates or may act as a negative regulator.