Arginine kinase (AK) plays an important role in the cellular energy metabolism of invertebrates. We investigated the effects of Zn2+ on Exopalaemon carinicauda arginine kinase (ECAK). Zn2+ conspicuously inactivated the activity of ECAK (IC50 = 8.49 +/- 0.76 mu M), and the double-reciprocal kinetics indicated that Zn2+ induced non-competitive inhibition of arginine and ATP. Spectrofluorometry results showed that Zn2+ induced tertiary structure changes in ECAK with the exposure of hydrophobic surfaces that directly induced ECAK aggregation. The addition of osmolytes, such as glycine and proline, successfully blocked the ECAK aggregation and restored the conformation and activity of ECAK. We measured the ORF gene sequence of ECAK using RACE and built a 3D structure of ECAK using homology models. Additionally, molecular dynamics (MD) and docking simulations between ECAK and Zn2+ have been conducted. The simulation results showed that Zn2+ blocked the entrance of ATP to the active site, and this result is consistent with.the experimental result showing Zn2+-induced inactivation of ECAK. Our study demonstrates the effect of Zn2+ on ECAK enzymatic function and unfolding, including aggregation, and the protective effects of osmolytes on ECAK folding. This study might provide important insights into the AK metabolic enzyme of invertebrates in marine environments. (C) 2014 Elsevier Ltd. All rights reserved.