ACETYLCHOLINE BINDING PROTEIN & ACETYLCHOLINE ESTERASE

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。我们已经通过合成一个与蜗牛结合蛋白同源的基因来制备烟碱型乙酰胆碱受体的胞外区。它显示了适当的配体专一性和流体力学性质，但我们希望检查它是否通过六聚体组装为环状五聚体而不是线性四聚体。几年前，埃利斯曼博士和我的实验室使用选择性免疫金标记法对突触中的乙酰胆碱酯酶进行了定位。多年来，定位工具有了很大的改进，我们现在有了一种小肽，束蛋白，它可以在低PM浓度下与乙酰胆碱酯酶相互作用。埃利斯曼实验室开发了一种新的曙红标记技术，大大提高了电子密度材料沉积的分辨率。因此，我们希望利用曙红标记的束蛋白来用更高分辨率的化学方法来定位突触的乙酰胆碱酯酶。我们的实验室有制作标签曙光的材料和技术

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. We have have prepared the extracellular domain of the nicotinic acetylcholine receptor by synthesizing a gene homologous with the snail binding protein. It shows the appropriate ligand specificity and hydrodynamic properties, but we would like to check to see whether it assembles as a circular pentamer rather than a linear tetramer through hexamer. Negative staining of the protein on a grid may resolve this issue.Part BSome years ago Dr Ellisman's and my laboratory employed selective immunogold labeling to localize acetylcholinesterase in the synapse. The tools for localization have greatly improved aver the years, and we now have a small peptide, fasciculin, that interacts with acetylcholinesterase at low pM concentrations. The Ellisman laboratory has developed a novel eosin labeling technique that greatly improves the resolution of deposition of electron dense materal. Hence, we hope to employ eosin-labeled fasciculin to localize synaptic acetylcholinesterase with higher resolution chemistry. Our laboratory has the materials and skills to make labeled eosi