Distinctive expression of a zinc-binding protein in rice callus grown in medium with high zinc concentration

Abstract

We investigated the growth, contents of water-soluble protein and free amino acids of the callus of japonica rice (Oryza sativa L. cv. Nipponbare) cultured in liquid N6 medium containing a high concentration of zinc. Furthermore, we determined the N-terminal amino acid sequence of a Zn-binding protein expressed in large quantities in the callus. The addition of Zn stimulated the growth of the callus and increased the Zn concentration. The callus subjected to the high-Zn treatment (hereafter referred to as CHZn) contained a larger amount of soluble proteins and a smaller amount of free amino acids than the control callus. Zinc-binding proteins were separated by affinity chromatography. The SDS-PAGE pattern of these proteins showed a distinctive protein band of about 29 kDa. Especially, CHZn contained larger quantities of 29 kDa protein than the control callus. Twenty-seven N-terminal amino acids of the protein were sequenced as DYAPMTLTIVNNCPYPVWPGIQANSGH. Results of homology search to the amino acid sequences from the nr-aa database and the dbEST database suggested that this 29 kDa protein may be a novel zinc-binding protein and that the protein may regulate the concentration of free zinc in the cytoplasm of callus cells through its binding to zinc ions.