Isolation and characterization of new isoforms of trichosanthin from Trichosanthes kirilowii

Three ribosome- inactivating proteins (RIPs) were isolated from the previously characterized 'trichosanthin' fraction prepared from the root tuber of Trichosanthes kirilowii. They were designated as alpha-, beta-, and gamma-trichosanthin (TCS). All three trichosanthin isoforms exhibited similar biochemical activities, namely in vitro inhibition of protein synthesis, N-glycosidase activity on 28S rRNA, and conversion of supercoiled DNA to open-circular and linear DNA. SDS-PAGE analysis indicated that their molecular weights were around 26 kDa. Amino acid composition analysis revealed that alpha-TCS. the major protein, corresponded closely to the previously reported trichosanthin. Protein finger-printing analysis indicated that gamma-TCS is structurally distinct from a-. and beta-TCS while the latter two are very similar, The identity of gamma-TCS was confirmed by N-terminal and internal protein sequencing. Cytotoxicity assay indicated that alpha-TCS is distinctively more toxic towards lymphocytes than beta- and gamma-TCS, while for fibroblast, beta-TCS is the least toxic. Their difference in cytotoxicity in spite of their structural similar warrant further investigation into their structure and function relationship. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved.