3 DISTINCT DOMAINS IN THE CHOLINESTERASE MOLECULE CONFER SELECTIVITY FOR ACETYLCHOLINESTERASE AND BUTYRYLCHOLINESTERASE INHIBITORS
3 DISTINCT DOMAINS IN THE CHOLINESTERASE MOLECULE CONFER SELECTIVITY FOR ACETYLCHOLINESTERASE AND BUTYRYLCHOLINESTERASE INHIBITORS
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DOI：
10.1021/bi00096a018
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发表时间：
1993-11-16
期刊：
影响因子：
2.9
通讯作者：
TAYLOR, P
中科院分区：
文献类型：
--
作者：
RADIC, Z;PICKERING, NA;TAYLOR, P
通过检查抑制剂与小鼠乙酰胆碱酯酶的单个和多个位点特异性突变体的相互作用，我们在胆碱酯酶结构中确定了三个不同的结构域，它们负责赋予乙酰胆碱酯酶抑制剂和丁酰胆碱酯酶抑制剂选择性。第一个域是最明显的；它定义了对酰基口袋尺寸的限制，其中 F295 和 F297 的侧链主要勾勒出乙酰胆碱酯酶中的该区域。用丁酰胆碱酯酶中发现的脂肪族残基替换这些苯丙氨酸侧链，可以催化更大的底物，并适应丁酰胆碱酯酶选择性烷基磷酸盐，例如 isoOMPA。此外，丁酰胆碱酯酶的底物激活特征元素在 F297I 突变体中也很明显。用酪氨酸取代 F295 和 F297 进一步改变了催化常数。第二个域位于活动中心峡谷边缘附近，由两个酪氨酸 Y72 和 Y124 以及 W286 定义；该区域似乎对于双四元抑制剂（例如 BW284C51）的选择性至关重要。第三个结构域定义了胆碱结合位点。在此，除了保守的 E202 和 W86 之外，仅在乙酰胆碱酯酶中发现的关键酪氨酸 Y337 负责空间封闭取代的三环抑制剂（例如乙丙嗪）的结合位点。对一系列取代吖啶和吩噻嗪的分析定义了配体上的基团和该位点控制结合选择性的氨基酸侧链。三个结构域中的每一个均由芳香族残基簇定义。稳定季铵部分的两个域各自含有负电荷，这有助于各自配合物的稳定能量。
By examining inhibitor interactions with single and multiple site-specific mutants of mouse acetylcholinesterase, we have identified three distinct domains in the cholinesterase structure that are responsible for conferring selectivity for acetyl- and butyrylcholinesterase inhibitors. The first domain is the most obvious; it defines the constraints on the acyl pocket dimensions where the side chains of F295 and F297 primarily outline this region in acetylcholinesterase. Replacement of these phenylalanine side chains with the aliphatic residues found in butyrylcholinesterase allows for the catalysis of larger substrates and accommodates butyrylcholinesterase-selective alkyl phosphates such as isoOMPA. Also, elements of substrate activation characteristic of butyrylcholinesterase are evident in the F297I mutant. Substitution of tyrosines for F295 and F297 further alters the catalytic constants. The second domain is found near the lip of the active center gorge defined by two tyrosines, Y72 and Y124, and by W286; this region appears to be critical for the selectivity of bisquaternary inhibitors, such as BW284C51. The third domain defines the site of choline binding. Herein, in addition to conserved E202 and W86, a critical tyrosine, Y337, found only in the acetylcholinesterases is responsible for sterically occluding the binding site for substituted tricyclic inhibitors such as ethopropazine. Analysis of a series of substituted acridines and phenothiazines defines the groups on the ligand and amino acid side chains in this site governing binding selectivity. Each of the three domains is defined by a cluster of aromatic residues. The two domains stabilizing the quaternary ammonium moieties each contain a negative charge, which contributes to the stabilization energy of the respective complexes.