Comparative Study Of Structure And Function Of The Lectin Subunits Of Cinphorin And Cinnamomin

Cinnamomin is a type II ribosome-inactivating protein (RIP) from the mature seeds of camphor tree (Cinnamomum camphora). It has three highly homologous isoforms, cinnamomin I, II, III, which appear as one band on the denatured polyacrylamide gel. Recently, a miniature type II RIP, cinphorin, was discovered in the seeds of camphor trees. Compared with cinnamomin, the A-chain of cinphorin was truncated and the B-chains of both cinnamomin and cinphorin appeared similar on polyacrylamide gel electrophoresis. It was also found that both A and B chain of the two toxins/lectins had the entirely identical N-terminal 10 amino acid sequence. It is already known that the B-chains of type II RIPs are lectins, which play an important role in the cytotoxicity of these proteins because binding of the B chain to sugar-containing receptors on the cell surface is a prerequisite for the toxic proteins to enter the cell. The intrinsic toxicity of type II RIP is apparently dependent to a great extent on the carbohydrate-binding activity and specificity of the lectin, since type I and type III RIP that have no B-chain fail to exhibit any cytotoxicity. In the thesis, the partial sequence of cinphorin B-chain was determined by LC/MS/MS and compared with that of cinnamomin. The results of MS/MS sequencing and amino acid composition analysis manifested that the primary structure of cinphorin B-chain was same as that of cinnamomin. The lectin activities of the two toxins were also investigated. They showed similar hemagglutination activity. Their saccharide binding specificities were studied by hapten inhibition and both of them were galactose-specific. However, N-acetylgalactosamine failed to bind to the two RIPs as ricin/abrin did. The interactions of the two lectins with specific saccharides were also investigated by fluorescence spectroscopy by which association constants were obtained. The association constants of galactose or lactose to them were found to be identical. These results indicated that the lectin subunit of cinphorin was identical with that of cinnamomin in both structure and function.