Active site of arginine kinase from sea cucumber Stichopus japonicus by site mutagenesis

Three mutations: C²⁷⁴A, R²⁸³G and H²⁸⁷A, of sea cucumber Stichopus japonicus dimeric arginine kinase were obtained by site mutagenesis to research the energy metabolism of invertebrate animals. The three mutants were then expressed in E. colt. The bulk of the expressed protein resided in insoluble inclusion bodies. The catalytic activity and structural features of the proteins were analyzed to show that all three mutants lost most of their catalytic activity. The mutant C²⁷⁴A lost almost all of its catalytic activity, but its structure was almost the same as the wild type. The results show that all three residues play important roles in maintaining either the structural or functional integrity of the arginine kinase. The Cys²⁷⁴ might be the active site of ariginine kinase.