THE BINDING-SITE OF THE NICOTINIC ACETYLCHOLINE-RECEPTOR IN ANIMAL SPECIES RESISTANT TO ALPHA-BUNGAROTOXIN
THE BINDING-SITE OF THE NICOTINIC ACETYLCHOLINE-RECEPTOR IN ANIMAL SPECIES RESISTANT TO ALPHA-BUNGAROTOXIN
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DOI：
10.1021/bi00028a029
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发表时间：
1995-07-18
期刊：
影响因子：
2.9
通讯作者：
FUCHS, S
中科院分区：
文献类型：
--
作者：
BARCHAN, D;OVADIA, M;FUCHS, S
烟碱型乙酰胆碱受体(AChR)的配体结合部位位于或-亚基中，在192和193位含有串联半胱氨酸。我们一直在分析几种对α-银环蛇毒素(α-BTX)表现出不同程度抗药性的动物的AChRs结合部位结构域。我们早期对蛇和猫鼬AChR的研究表明，AChRα亚单位187、189和194位的氨基酸替换在确定这些ACNR对α-BTX的抗性方面起着重要作用。在本研究中，我们研究了刺猬、地鼠、猫和人的AChR结合部位结构与α-BTX结合的相关性。克隆了与这些物种的122-205位氨基酸残基相对应的AChRα亚基片段，对其进行了测序，并在大肠杆菌中表达。刺猬片段不与α-BTX结合，与蛇和猫鼬的AChR一样，而人片段是部分结合蛋白。地鼠片段和猫片段与α-BTX结合的程度与小鼠片段相似。刺猬和人的AChRs在阿尔法亚基的187和189位有非芳香族氨基酸残基，与“抗毒素”的蛇和猫鼬一样，与在这两个位置有芳香残基的“毒素粘合剂”形成对比。因此，ACnCα亚基187和189位的芳香族氨基酸残基似乎是与α-BTX正确结合所必需的，并且这些位置上的非芳香族残基的变化对于确定对α-BTX的抗性是重要的。
The ligand binding site of the nicotinic acetylcholine receptor (AChR) is located in the or-subunit, within a small fragment containing the tandem cysteines at positions 192 and 193. We have been analyzing the binding site domain of AChRs from several animal species exhibiting various degrees of resistance to alpha-bungarotoxin (alpha-BTX). Our earlier work on the snake and mongoose AChR, both of which do not bind alpha-BTX, suggested that amino acid substitutions at positions 187, 189, and 194 of the AChR alpha-subunit are important in determining the resistance of these ACNRs to alpha-BTX. In the present study, we have examined the correlation between alpha-BTX binding and the structure of the binding site domain of AChR from the hedgehog, shrew, cat, and human. Fragments of the AChR alpha-subunit corresponding to residues 122-205 from these species were cloned, sequenced, and expressed in Escherichia coli. The hedgehog fragment does not bind alpha-BTX, in common with the snake and mongoose AChR, and the human fragment is a partial binder. The shrew and cat fragments bind alpha-BTX to a similar extent as the mouse fragment. The hedgehog and human AChRs have nonaromatic amino acid residues at positions 187 and 189 of the alpha-subunit, as is seen with the ''toxin resistant'' snake and mongoose, and in contrast with the ''toxin binders'', which have aromatic residues at these two positions. Thus, it appears that aromatic amino acid residues at positions 187 and 189 of the ACNC alpha-subunit are required for proper alpha-BTX binding, and that changes at these positions to nonaromatic residues are important in determining resistance to alpha-BTX.