STRUCTURAL STUDIES OF ACETYLCHOLINE BINDING SITE

乙酰胆碱受体（AChR）是一种大的膜结合受体， 300 kDalton大小的蛋白质复合物，形成钠离子通道， 神经和肌肉细胞之间的连接处。 它的结构只是 已知9埃分辨率。 因此，没有原子分辨率的结构细节可用。 α-银环蛇毒素（BGTX），一种来自蛇毒液的强效神经毒素， 金环蛇，是一种非常有效的抑制剂， AChR，阻断乙酰胆碱结合位点。 好地完成了 BGTX的NMR溶液结构的精细化。 最小的 与BGTX强烈结合的AChR肽片段是12个残基 对应于以下α-亚基的残基185-196的片段 乙酰胆碱受体 我们已经能够观察和分配大部分的1H BGTX和AChR的12个残基片段的复合物中的共振， 并且计算出的溶液结构与核磁共振结果一致 限制，我们已经公布了。 互动细节 在该肽和BGTX之间包括产生疏水口袋 在12聚体肽的瓦尔-188、Tyr-190和缬氨酸39和40之间 关于BGTX 我们也一直致力于构象的研究 AChR的五个残基片段， 半胱氨酸（192和193），其在 vivo. 这些半胱氨酸已被证明是必不可少的， 通道开放，并在8埃内的硼和乙酰胆碱。 我们已经能够确定这种肽的三种构象， 其中Cys-192和Cys-193之间的酰胺键处于顺式 一个是反式构象，另一个是反式构象。 从最低能量的顺式构象到 其它顺式构象约为15千卡/摩尔，而势垒 从顺式到反式的转化率约为20 千卡/摩尔。 这些障碍与 从静息状态到AChR的脱敏状态， 这些八元环的构象过程 在顺序的二硫化物192和193之间可以 负责体内受体的脱敏。 我们 我已经完成了理论计算使用琥珀，我们是在 把这些结果写出来发表的过程。 计算机 图形实验室在分析我们的结果时是必不可少的。 这个重要的八元环的构象建模 含有肽。

The acetylcholine receptor (AChR) is a large membrane bound protein complex of 300 kDalton size, forming a sodium ion channel at the junction between nerve and muscle cells. Its structure is only known to 9 Angstroms resolution. Thus no details of the structure at atomic resolution are available. alpha-Bungarotoxin (BGTX), a potent neurotoxin from the venom of the banded krait Bungarus Multicinctus, is a very potent inhibitor of AChR, blocking the acetylcholine binding site. We have completed the refinement of the NMR solution structure of BGTX. The smallest peptide fragment of AChR that binds strongly to BGTX is a 12-residue fragment corresponding to residues 185-196 of the alpha-subunit of AChR. We have been able to observe and assign most of the 1H resonances in the complex of BGTX and the 12-residue fragment of AChR, and have calculated solution structures consistent with the NMR constraints, which we have published. Details of the interaction between this peptide and BGTX include a hydrophobic pocket is created between val-188, tyr-190 of the 12mer peptide, and valines 39 and 40 of BGTX. We have also been working on the study of the conformations of a five residue fragment of AChR containing the two sequential cysteines (192 and 193) which are disulfide-linked to each other in vivo. These cysteines have been demonstrated to be essential for channel opening, and are within 8 Angstroms of bo und acetylcholine. We have been able to identify three conformations of this peptide, two with the amide bond between cys-192 and cys-193 in the cis conformation, and one with this amide bond in the trans conformation. The barrier in going from the lowest energy cis conformation to the other cis conformation is approximately 15 kcal/mole, and the barrier in going from the cis forms to the trans form is approximately 20 kcal/mole. These barriers correlate well with the barriers in going from the resting state to the desensitized state of the AChR, so that these conformational processes of the eight-membered ring formed between the sequential disulfides 192 and 193 may be responsible for the desensitization of the receptor in vivo. We have completed theoretical calculations using AMBER, and we are in the process of writing up these results for publication. The Computer Graphics Labroatory is essential in analyzing the results of our modeling of the conformations of this important eight-membered ring containing peptide.