Ludueña R F, Roach M C, Prasad V, Pettit G R
Department of Biochemistry, University of Texas Health Science Center, San Antonio 78284-7760.
Biochem Pharmacol. 1993 Jan 26;45(2):421-7. doi: 10.1016/0006-2952(93)90079-c.
Halichondrin B is a polyether macrolide of marine origin which binds to tubulin and inhibits microtubule assembly in vitro and in vivo. As is the case with phomopsin A and dolastatin 10, halichondrin B is a non-competitive inhibitor of vinblastine binding to tubulin. Analogous to maytansine, which by contrast is a competitive inhibitor of vinblastine binding, halichondrin B has no effect on colchicine binding, which is greatly stabilized by phomopsin A and dolastatin 10, but not by maytansine. We have previously developed assays which allow sensitive discrimination among the interactions of various ligands with tubulin, and examined the effects of ligands on the reactivity of tubulin sulfhydryl groups and the exposure of hydrophobic areas on the surface of the tubulin molecule. To classify the nature of the interaction between halichondrin B and tubulin, in this study we examined the effects of halichondrin B and its closely related analogue, homohalichondrin B, by these assays. We found that: (1) halichondrin B and homohalichondrin B both inhibited formation of an intra-chain cross-link between two sulfhydryl groups in beta-tubulin, as do phomopsin A, dolastatin 10, maytansine, and vinblastine; (2) halichondrin B resembles maytansine in that it had no effect on alkylation of tubulin sulfhydryl groups by iodoacetamide, unlike phomopsin A, dolastatin 10 and vinblastine, all of which inhibit alkylation; (3) halichondrin B differs from other anti-mitotic drugs in that it enhanced exposure of hydrophobic areas on tubulin; (4) homohalichondrin B, like maytansine and in contrast to phomopsin A, dolastatin 10 and vinblastine, had no effect on exposure of hydrophobic areas; and (5) homohalichondrin B, contrary to maytansine, inhibited alkylation of tubulin sulfhydryl groups in the presence of GTP and MgCl2. In their interactions with the tubulin molecule, halichondrin B and homohalichondrin B appear to have unique conformational effects which differ from those of other drugs and also from the effects of each other as well.
海兔毒素B是一种源自海洋的聚醚大环内酯，它能与微管蛋白结合，并在体内外抑制微管组装。与腐皮镰刀菌毒素A和多拉司他汀10的情况一样，海兔毒素B是长春碱与微管蛋白结合的非竞争性抑制剂。与美登素相反，美登素是长春碱结合的竞争性抑制剂，海兔毒素B对秋水仙碱结合没有影响，秋水仙碱结合在腐皮镰刀菌毒素A和多拉司他汀10作用下能得到极大稳定，但在美登素作用下则不然。我们之前开发了一些检测方法，可灵敏地区分各种配体与微管蛋白的相互作用，并研究了配体对微管蛋白巯基反应性以及微管蛋白分子表面疏水区域暴露的影响。为了对海兔毒素B与微管蛋白之间相互作用的性质进行分类，在本研究中我们通过这些检测方法研究了海兔毒素B及其密切相关类似物高海兔毒素B的作用。我们发现：（1）海兔毒素B和高海兔毒素B均抑制β-微管蛋白中两个巯基之间链内交联的形成，腐皮镰刀菌毒素A、多拉司他汀10、美登素和长春碱也有同样作用；（2）海兔毒素B与美登素类似，对碘乙酰胺对微管蛋白巯基的烷基化没有影响，这与腐皮镰刀菌毒素A、多拉司他汀10和长春碱不同，它们均抑制烷基化；（3）海兔毒素B与其他抗有丝分裂药物不同，它能增强微管蛋白上疏水区域的暴露；（4）高海兔毒素B与美登素一样，与腐皮镰刀菌毒素A、多拉司他汀10和长春碱相反，对疏水区域的暴露没有影响；（5）与美登素相反，高海兔毒素B在存在鸟苷三磷酸（GTP）和氯化镁（MgCl2）的情况下抑制微管蛋白巯基的烷基化。在与微管蛋白分子的相互作用中，海兔毒素B和高海兔毒素B似乎具有独特的构象效应，这些效应不同于其他药物，彼此之间也不相同。
