Reconsideration of the catalytic center and mechanism of mammalian paraoxonase/arylesterase.
Reconsideration of the catalytic center and mechanism of mammalian paraoxonase/arylesterase.
复制标题
哺乳动物对氧磷酶/芳基酯酶催化中心和机制的重新思考。
DOI：
10.1073/pnas.92.16.7187
复制
发表时间：
1995
影响因子：
11.1
通讯作者：
LaDu,BN
中科院分区：
文献类型：
--
作者：
Sorenson,RC;Primo-Parmo,SL;Kuo,CL;Adkins,S;Lockridge,O;LaDu,BN
三十年来，哺乳动物对氧磷酶（A-酯酶、芳香族酯酶、芳基酯酶; PON，EC 3.1.8.1）被认为是半胱氨酸酯酶，证明与丝氨酸酯酶（胆碱酯酶和羧酸酯酶）具有结构和机制同源性。人、小鼠和兔的脑桥各仅含有三个半胱氨酸残基，并且它们在PON中的位置是保守的。在纯化的人PON中，残基Cys-41和Cys-352形成分子内二硫键，两者都不能作为活性中心半胱氨酸发挥作用。高度纯化的酶活性PON含有单个可滴定的巯基。因此，Cys-283是活性中心半胱氨酸的唯一可能候选者。通过对人cDNA的定点突变，Cys-283被丝氨酸（C283 S）或丙氨酸（C283 A）取代。表达的C283（野生型）酶被对羟基苯甲酸汞灭活，但C283 S和C283 A突变体酶未被灭活。C283 A和C283 S突变酶保留对氧磷酶和芳基酯酶活性，对氧磷和乙酸苯酯的Km值与野生型相似。显然，残基Cys-283在活性PON中是游离的，但对氧磷酶或芳基酯酶活性不需要游离巯基。因此，有必要研究PON的活性中心结构和催化机制的其他模型。
For three decades, mammalian paraoxonase (A-esterase, aromatic esterase, arylesterase; PON, EC 3.1.8.1) has been thought to be a cysteine esterase demonstrating structural and mechanistic homologies with the serine esterases (cholinesterases and carboxyesterases). Human, mouse, and rabbit PONs each contain only three cysteine residues, and their positions within PON have been conserved. In purified human PON, residues Cys-41 and Cys-352 form an intramolecular disulfide bond and neither could function as an active-center cysteine. Highly purified, enzymatically active PON contains a single titratable sulfhydryl group. Thus, Cys-283 is the only probable candidate for an active-center cysteine. Through site-directed mutagenesis of the human cDNA, Cys-283 was replaced with either serine (C283S) or alanine (C283A). The expressed C283 (wild-type) enzyme was inactivated by para-hydroxymercuribenzoate, but the C283S and C283A mutant enzymes were not inactivated. C283A and C283S mutant enzymes retained both paraoxonase and arylesterase activities, and the Km values for paraoxon and phenyl acetate were similar to those of the wild type. Clearly, residue Cys-283 is free in active PON, but a free sulfhydryl group is not required for either paraoxonase or arylesterase activities. Consequently, it is necessary to examine other models for the active-site structure and catalytic mechanism of PON.
人血清对氧磷酶/芳基酯酶的遗传决定的同种酶形式的特征。
DOI：
--
发表时间：
1991
期刊：
Drug metabolism and disposition: the biological fate of chemicals
影响因子：
--
作者：
Smolen,A;Eckerson,HW;Gan,KN;Hailat,N;LaDu,BN
通讯作者：
LaDu,BN