Microtubule regulation by small molecules.

在第一部分中，我们将重点放在抗有丝分裂肽上，因为它们是已知的最有效的抗MT药物之一，它们已经被合成并且有类似物可用，并且因为它们诱导MT亚基呈现不寻常的特征环状。我们正在研究这些环状聚合物的结构和动力学性质，通过分析超离心，低温电子显微镜，荧光相关光谱，和蛋白酶作图。我们的研究揭示了这些环的高稳定性和均匀性，这促使我们尝试这些聚合物的结晶，以实现其结构的原子分辨率。我们还研究了沙利度胺和康布他汀a的合成类似物对微管聚合的影响。我们还证明了去乙酰化酶翻译后修饰的作用，以及抗微管药物对p53反应的影响。

In part one we have focused on antimitotic peptides because these are among the most potent anti-MT agents known, they have been synthesized and analogs are available, and because they induce the MT subunits to assume unusual and characteristic ring shapes. We are studying the structural and dynamic properties of these ring polymers by analytical ultracentrifugation, cryoelectron microscopy, fluorescence correlation spectroscopy, and protease mapping. The high stability and uniformity of these rings that our studies revealed have led us to attempt crystallization of these polymers to achieve atomic resolution of their structure. We are also examining the effects on microtubule polymerization of synthetic analogs of thalidomide and combretastatin A. We have also demonstrated the role of posttranslational modifications by deacetylases, and the effects of antimicrotubule drugs on the p53 response. In part two, we are seeking to identify small molecules that do not bind well with mammalian tubulin but do bind to parasite tubulin. The tubulin molecule is quite conserved evolutionarily, but differences do exist, and several molecules are known that can target, for example, yeast rather than mammalian tubulin or vice-versa. We are looking for molecules that will target Leishmania, the infectious cause of an important group of human diseases. We have identified several small molecules that show promise as selective agents, binding to Leishmania tubulin preferentially over mammalian tubulin, and preventing parasite multiplication inside human macrophage cells. In addition, we have produced purified tubulin from cultured Leishmania and evaluated it for use in drug screening.