The Interaction Of Metal Ions And Trichosanthin

The structure and function of metalloproteins and metalloenzymes have been the center of the attention for many years due to their important roles in biological system.Though metal ions exist in trace amount in metalloproteins and metalloenzymes, their roles in structure or catalysis are crucial. The metal ions and its coordination environment are generally the active center of the biological molecules, so the study on the metal ions and their micro-environment is of great significance.Trichosanthin (TCS) is a single-chain ribosome inactivating protein. TCS belongs to a sort of metalloprotein, and Ca2+ locates in its active center. It is isolated from Trichosanthes kirilowii tubers of cucurbitaceae and is the active component of Chinese drug Tian Hua Fen. Recent studies indicated that TCS has curative effect for the treatment of cancer, AIDS etc. Therefore, studies on TCS are of important values in both academic research and practical application.This study focused on the interaction of TCS with some important essential metal ions in organism, such as Zn2+, Cu2+, Co2+, Ca2+, Fe3+, by fluorescence spectrum and UV spectrum. At the same time, the influence of metal ions on the structure of TCS was also studied.Following are the key experimental results obtained:At physiologically-relevant conditions (i.e. Britton-Robinson buffer solution at pH=6.8, normal saline), all the metal ions examined including Co2+, Ca2+, Zn2+, Cu2+, Fe3+ (in the concentration range of 0~0.5mmol/L) do not shift the maximum emission band of TCS. Among them, Co2+ and Ca2+ ions have no obvious effect on endogenesis fluorescence intensity of TCS, and Zn2+ has some weak effects.However, Cu2+ and Fe3+ have obvious quenching effect, although their quenching process is different, with Fe3+ being more pronounced than Cu2+.The change of pH could not revert the Fe3t induced fluorescent quenching of TCS. However, the emission spectra at 295nm were different after the addition of Fe3+.The interaction of TCS with EDTA was also studied in this work, which showed that EDTA have weak effect on endogenesis Fluorescence of TCS even at concentration as high as 40mmol/L ( [EDTA]/[TCS]=2.00 104).In conclusion, the five metal ions examined do not shift the maximum fluorescent emission at 332nm, while Cu2+and Fe3+ can disturb the structure of TCS and perhaps affect the function of protein in some extent. Though Zn 2+ and Co 2+ are divalent ions, they probably can not substitute Ca2+ from the active center of TCS, or can substitute Ca2+but does not change the structure of TCS. There is no significant change observed for the fluorescence intensity of TCS. High concentration of Ca2+ also cannot affect the structure of TCS.