STRUCTURAL BASIS FOR MICROTUBULE NUCLEATION BY THE GAMMA-TUBULIN SMALL COMPLEX

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 中心体在组织微管细胞骨架中起着基础性作用。中心体功能缺陷会导致染色体分离错误，这是癌症发生和发展的主要因素。我们的目标是通过研究所有真核生物中一个关键的300 kDa异四聚体复合体的结构来了解中心体微管成核的机制和调控：β-微管蛋白小复合体(？-TUSC)。我们用随机圆锥倾斜法测定了负应变的EM结构，并在低温条件下得到了初步的图像。从更高分辨率的冷冻EM结构中，我们希望确定复合体的整体组织，特别是微管负端与微管蛋白相互作用的微管蛋白表面，并绘制连接微管组织中心的蛋白质的结合位置。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The centrosome plays a fundamental role in organizing the microtubule cytoskeleton. Defects in centrosome function lead to errors in chromosome segregation, a major factor in both the initiation and progression of cancer. We aim to understand the mechanism and regulation of microtubule nucleation at the centrosome by focusing on the structure of a key 300 kDa heterotetrameric complex that underlies nucleation in all eukaryotes: the ¿-tubulin small complex (¿-TuSC). We have determined a negative stain EM structure by the random conical tilt method, and have preliminary images under cryo conditions. From a higher resolution cryo-EM structure we hope to determine the overall organization of the complex and particularly which surfaces of ¿-tubulin are exposed for interaction with ¿¿-tubulin at the microtubule minus ends, and to map the binding sites of proteins which link ¿-TuSC to microtubule organizing centers.