Jatropha (jatropha Curcas) Ribosome-inactivating Proteins (curcin) Isolated And Purified

Ribosome-inactivating protein (RIPs) of plant is known for its biological activities such as anti-tumor, anti-birth, anti-HIV virus, anti-parasite, antifungal and insecticidal activities and its wide application in anti-virus genetic engineering of plant. Therefore, RIPs has enticing prospect in application for its various activities. However, few reports on purification and biological activity of curcin have been reported in the literatures. In view of these facts, the purification conditions were optimized and bioactivity of curcin was evaluated in the present work.The contents of total protein, ash and moisture from fresh seeds of Jatropha curcas Linna in Guizhou province were determined and found to be 24.17%, 8.00% and 4.40%, respectively. Moreover, total protein content from fresh seeds of Jatropha curcas Linna in Guizhou province was found to be higher than that from stored seeds.Two proteins were obtained from kernels of Jatropha curcas Linna with 95% saturation of (NH₄)₂SO₄ and by the gel filtration chromatography on sephadex G-100. One protein was identified as curcin (the toxalbumin), a ribosome-inactivating protein, whose relative molecular weight and isoelectric point were 29.10 kDa and 8.70, respectively. The other one was a small molecular protein with relative molecular weight 10kDa.The orthogonal experiment was studied with some variable parameters such as the concentration of phosphate buffer (PBS), pH value and the concentration of NaCl solution. Proteins were extracted from kernels of Jatropha curcas Linna with phosphate buffer of different concentration. In order to seek the optimal extraction conditions of crude proteins from Jatropha curcas Linna kernels, the protein patterns of SDS-PAGE were analyzed with the software of quantity one, and the percentage of curcin was tested. The highest extraction percentage of 0.103% for curcin was achieved with 0.05 mol/L phosphate buffer (pH = 6.80) containing 0.14 mol/L NaCl. In addition, the condition for purification of crude proteins from Jatropha curcas Linna kernels was selected. The result showed that two different proteins could be separated well when the height of the column used in chromatography was 80 cm. Proteins were evaluated for anticancer activity in vitro. The results of bioassay indicated that curcin had no inhibition effect on PC3, BGC823 cells, but showed some effect against Bcap37 cell. The other protein (10 kDa) had no inhibition effect on PC3, BGC823 and Bcap37 cells.