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

• MD simulations for the action of Pb2+ on the inhibition of alpha-glucosidase.
• The predicted docking residues for Pb2+ in the active site of alpha-glucosidase.
• The inhibitory effect and binding mechanisms of Pb2+ on alpha-glucosidase.
• Structural changes of alpha-glucosidase induced by Pb2+.
• Integrating study between inhibition kinetics and computational simulations.

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 Pb2+. As a result, the binding mechanism of Pb2+ to ECAK was predicted from the binding sites, and the structural change was elucidated. To confirm the simulated results, the Pb2+-mediated inhibition and aggregation of ECAK were subsequently conducted. We found that Pb2+ partially inactivated the activity of ECAK with relatively strong binding. The spectrofluorimetry results showed that Pb2+ induced tertiary structural changes of ECAK, with the substantial exposure of hydrophobic surfaces directly induced by ECAK aggregation. The ECAK aggregation process induced by Pb2+ occurred with multi-phase kinetics. The addition of osmolytes did not protect ECAK from Pb2+ inactivation. Because AK plays an important role in the cellular energy metabolism of invertebrates, our study suggests new information about the effect of Pb2+ on ECAK's enzymatic function and unfolding, including aggregation, which may be toxic to invertebrates or may act as a negative regulator.

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