MEMBRANE-SPANNING DOMAINS IN ACETYCHOLINE RECEPTOR

乙酰胆碱受体是一种跨越质膜和 在细胞质和胞外之间形成阳离子的门控通道 肌肉细胞。这条航道中通过 膜本身由五个环组成，并置，疏水 结构域，五个亚基中的每个亚基一个。这是我们的长期目标。 来确定每一个分子的精确分子结构 域，因此环本身和通道的域 阳离子。四个亚基中每个亚基中的四个跨膜序列 已经鉴定出乙酰胆碱受体，Kendrew-Watson导线 已经建立了四个跨膜螺旋的模型 伽玛亚单位。这四个山丘已经排成一条线，形成了一个 这一跨膜结构域的假想分子模型 蛋白。该模型包含两个亚基内的二硫键 双层的。这两个二硫化物将在本地结构中寻找 乙酰胆碱受体的化学实验 纯化的蛋白质。含有这些二硫化物的多肽将被分离出来 并由已成功用于提纯 (Na++K+)-ATPase的跨膜部分。如果这两个都是二硫化物 存在的情况下，假设模型很可能是正确的。 如果不这样做，可能会在 天然结构中其他亚基的跨膜结构域，以及 这些也将被寻求。如果11种独特的半胱氨酸中的每一种 然而，膜内的残留物是自由的，交联剂将 附着于可从烷基核接触到的任何一种 双层的。这些半胱氨酸与邻近化合物之间的共价交联键 会形成氨基酸侧链，这些侧链的产物 将对交联反应进行纯化和鉴定。如果一个或两个 上述化学实验中的任何一项都有阳性结果，它们 应该为跨膜结构域定义一个独特的分子结构 乙酰胆碱受体四个亚基中的至少一个。这, 但是，将定义四个域中每个域的结构 这四个亚基因为每个亚基都必须折叠成相同的， 可叠加的模式。

Acetylcholine receptor is a protein that spans the plasma membrane and forms a gated channel for cations between the cytoplasm and the exterior of the muscle cell. The portion of this channel that passes through the membrane itself is formed by a ring of five, juxtaposed, hydrophobic domains, one from each of the five subunits. It is the long-term objective of this project to define the precise molecular structure of each of these domains and, thereby, that of the ring itself and the channel for the cations. The four membrane-spanning sequences in each of the four subunits of acetylcholine receptor have been identified, and Kendrew-Watson wire models have been constructed of the four membrane-spanning helices of the Gamma subunit. These four hilices have been aligned to create a hypothetical molecular model for the membrane-spanning domain of this protein. This model contains two intrasubunit disulfide bonds within the bilayer. These two disulfides will be sought within the native structure of acetylcholine receptor through chemical experiments performed on the purified protein. Peptides containing these disulfides will be isolated and identified by methods that have been used successfully to purify the membrane-spanning portions of (Na+ + K+)-ATPase. If these disulfides both exist, there is a high probability that the hypothetical model is correct. If they do not, other intrasubunit disulfides may occur within the membrane-spanning domains of other subunits in the native structure, and these will also be sought. If every one of the eleven unique cysteine residues within the membrane, however, are free, cross-linking agents will be attached to any of them that are accessible from the alkane core of the bilayer. Covalent cross-links between these cysteines and neighboring amino acid side chains will be formed, and the products of these cross-linking reactions will be purified and identified. If one or two of any of the foregoing chemical experiments provide positive results, they should define a unique molecular structure for the membrane-spanning domain of at least one of the four subunits of acetylcholine receptor. This, however, will define the structure of each of the four domains from each of the four subunits because every subunit is necessarily folded in the same, superimposable pattern.