γ-Glutamyl-cysteine synthetase from bovine lens: I. Purification and properties
γ-Glutamyl-cysteine synthetase from bovine lens: I. Purification and properties
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牛晶状体γ-谷氨酰半胱氨酸合成酶：一、纯化及性质
DOI：
10.1016/0003-9861(67)90124-5
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发表时间：
1967
影响因子：
3.9
通讯作者：
W. Rathbun
中科院分区：
文献类型：
--
作者：
W. Rathbun
催化谷胱甘肽合成第一步的牛晶状体酶γ-谷氨酰半胱氨酸合成酶被纯化了260倍，恢复了60%的原始活性。酶的不稳定性要求在接近其冰点时迅速使用每种溶剂。在分离的进展之后，用兔抗血清对两个纯化的部分进行免疫电泳。对于谷氨酸、ATP、半胱氨酸和α-氨基丁酸盐的明显km值测定，ADP的抑制作用需要外推初始速度。对谷氨酸部分的要求是一个五碳二羧酸，其构型中有一个游离氨基。异羟基谷氨酸、α-甲基谷氨酸和β-羟谷氨酸能够取代-谷氨酸。半胱氨酸部分需要这种构型；α-氨基丁酸盐取代半胱氨酸。来自牛晶状体和猪肝的酶在各种条件下的最佳pH值和不稳定性不同。这表明两种来源的酶可能是完全不同的蛋白质。
The bovine lens enzyme catalyzing the first step of glutathione synthesis, γ-glutamyl-cysteine synthetase, was purified 260-fold with a recovery of 60% of the original activity. The instability of the enzyme required the rapid use of each solvent near its freezing point. Progress in fractionation was followed by immunoelectrophoresis with rabbit antisera produced in response to two of the purified fractions. Inhibition by ADP required extrapolation to initial velocity for apparentKmvalus determinations of glutamate, ATP, cysteine, and α-aminobutyrate. The requirements for the glutamate moiety were a five-carbon dicarboxylic acid with a free amino group in thel-configuration. Allohydroxyglutamate, α-methylglutamate and β-hydroxyglutamate were capable of substituting forl-glutamate. Thel-configuratioii was required for the cysteine moiety; α-amino butyrate substituted forl-cysteine. The enzyme from bovine lens and hog liver differ in their pH optima and instability to a variety of conditions. This indicates the enzymes from the two sources may be distinctly different proteins.