Development of biotin-avidin technology to investigate okadaic acid-promoted cell signaling pathway
Development of biotin-avidin technology to investigate okadaic acid-promoted cell signaling pathway
复制标题
DOI：
10.1016/s0040-4020(00)00752-3
复制
发表时间：
2000-11-10
期刊：
影响因子：
2.1
通讯作者：
Hatanaka, Y
中科院分区：
文献类型：
--
作者：
Konoki, K;Sugiyama, N;Hatanaka, Y
合成了4种冈田酸生物素偶联物，用表面等离子体共振（SPR）研究了它们与蛋白磷酸酶2A（PP 2A）的相互作用。C7-生物素化冈田酸表现出最强的结合亲和力的酶，而Cl-生物素化衍生物是缺乏亲和力。C24-或C27-生物素化的冈田酸显示出对酶的中等亲和力。在这一发现之后，将生物素基光亲和探针引入到冈田酸的7-OH中。光亲和标记后的SDS-PAGE分析表明，冈田酸衍生物清楚地标记PP 2A。此外，还标记了三种蛋白质的海洋海绵Halichondria okadai的粗提物。所有这些结果表明，C7-生物素缀合物是一种通用的试剂，用于冈田酸结合蛋白，包括PP 2A的生化研究。(C)2000爱思唯尔科技有限公司版权所有。
Four biotin conjugates of okadaic acid were synthesized for evaluating their interactions with protein phosphatase 2A (PP2A) by surface plasmon resonance (SPR). C7-biotinylated okadaic acid exhibited the strongest binding affinity to the enzyme, while Cl-biotinylated derivative was devoid of affinity. C24- or C27-biotinylated okadaic acid showed moderate affinity to the enzyme. In the wake of this finding, a biotinyl photoaffinity probe was introduced into 7-OH of okadaic acid. Photoaffinity labeling followed by SDS-PAGE analysis indicated that the okadaic acid derivative clearly labeled PP2A. Furthermore, three proteins were also labeled in crude extracts of a marine sponge Halichondria okadai. All these results imply that the C7-biotin conjugate is a versatile reagent for biochemical studies of okadaic acid-binding proteins including PP2A. (C) 2000 Elsevier Science Ltd. All rights reserved.