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本文报道我室发现的一种天然钙调素拮抗剂马兜铃酸的研究。利用丹磺酰标记的 CaM(D-CaM)对马兜铃酸的研究表明,马兜铃酸是一种非钙离子依赖性钙调素拮抗剂。实验测得马兜铃酸与 D-CaM 结合的解离常数,有 Ca~(2+)和无 Ca~(2+)情况下分别为70μmol/L、77μmol/L。两种状况下马兜铃酸对 D-CaM 荧光强度的抑制分别为40%、41%。暗示马兜铃酸主要作用于 CaM 上 Ca~(2+)诱导的疏水区之外。三氟啦礤(TFP)引起的D-CaM 荧光增强可被马兜铃酸明显降低,而 TFP 在达到马兜铃酸浓度的15倍以上,仍未能递转马兜铃酸对D-CaM 荧光强度的降低作用,这为马兜铃酸主要作用于 CaM 上 Ca~(2+)诱导的疏水区以外提供了又一佐证。
This article reports the study of aristolochic acid, a natural calmodulin antagonist found in our laboratory. The study of aristolochic acid using dansyl labeled CaM (D-CaM) showed that aristolochic acid is a non-calcium-dependent calmodulin antagonist. The dissociation constants for the binding of aristolochic acid to D-CaM were determined experimentally and were found to be 70 μmol/L and 77 μmol/L in the presence of Ca 2+ and Ca 2+ . In both cases, the inhibition of D-CaM fluorescence intensity by aristolochic acid was 40% and 41%, respectively. It is suggested that aristolochic acid mainly acts on the Ca(2+) induced Ca(2+)-induced hydrophobic region. D-CaM fluorescence enhancement induced by trifluorophosphate (TFP) can be significantly reduced by aristolochic acid, while TFP can not deliver aristolochic acid to D-CaM when it reaches more than 15 times of aristolochic acid concentration. The decrease of fluorescence intensity provides further evidence that aristolochic acid mainly acts on the Ca 2+ -induced hydrophobic region outside CaM.