ENZYMOLOGY OF GOLGI STACK FORMATION

我们从小到大重建了高尔基堆积的形成 囊泡的平均直径为60-90 nm。我们发现了一种细胞质 含有120和66kD摩尔分子的两种蛋白质的组份。WT(p120/66)， 这对于组装过程的非常早期阶段是必需的。我们的 结果提示，这些蛋白质可能参与解开 来自细胞质锚的VGM。然后是自由的或移动的VGM 通过NSF和SNAP依赖的过程融合形成更大的囊泡 直径约200-300 nm。我们将正式测试p120/66的作用 在动员VGM将其组装成堆栈方面。此外，我们 提议确定p120/66互动伙伴。在……面前 一种名为p97的类似NSF的ATPase，大小泡融合并产生 膜被压扁成圆盘状的池子。我们建议P97 可能参与融合后较大小泡的扁平化。在……里面 此外，我们已经开始对外周高尔基体膜进行分级 蛋白质，并在三个柱层析步骤后找到一个组分 以遏制堆叠形成活动。我们将继续这样做 分级方案，以确定必要的成分。一种胞质 含有55和33kD多肽的蛋白质组分(P55/33) GTP的存在抑制了智商作用下高尔基体囊泡的形成。我们建议 这些蛋白质可能在高尔基体膜上交换GTP 蛋白。因此，对这些蛋白质的表征有望 揭示与高尔基体相关的GTP接受者，其结果是 抑制智商的影响。因此，我们的目标承诺揭示小说 对高尔基体堆叠形成机制的洞察 高尔基体可通过以下方式防止其完全囊泡形成的过程 利用GTP依赖反应。

We have reconstituted the formation of Golgi stack formation from small vesicles of 60-90 nm average diameter. We have identified a cytosolic fraction containing two proteins of 120 and 66 kD mol. wt (p120/66), which is required for very early stages of the assembly process. Our results suggest that these proteins may be involved in untethering of VGMs from a cytosolic anchor. The untethered or mobilized VGMs then fuse by an NSF and SNAP dependent process to form larger vesicles of about 200-300 nm diameter. We will formally test the role of p120/66 in mobilizing VGMs for their assembly into stacks. In addition, we propose to identify p120/66 interacting partners. In the presence of an NSF like ATPase called p97, the large vesicles fuse and the resulting membranes are flattened into disc shaped cisternae. We suggest the p97 may be involved in post fusion flattening of the larger vesicles. In addition, we have started fractionating peripheral Golgi membrane proteins and find a fraction, after three column chromatographic steps to contain the stack formation activity. We will continue with this fractionation scheme to identify the essential components. A cytosolic protein fraction containing polypeptides of 55 and 33 kD (p55/33) in the presence of GTP inhibits Golgi vesiculation by IQ. We propose that these proteins may be exchanging GTP onto a Golgi membranes associated protein. The characterization of these proteins, therefore, promises to reveal the Golgi associated recipient of GTP, which as a result inhibits the effects of IQ. Our aims therefore promise to reveal novel insights into the mechanism by which Golgi stacks form from VGMs and the process by which the Golgi may prevent its complete vesiculation by utilizing a GTP dependent reaction.