鱼类和哺乳动物中的抗坏血酸 -2- 硫酸酯硫酸酯酶。比较特征及在抗坏血酸代谢中的可能作用

Ascorbate-2-sulfate sulfohydrolase in fish and mammal. Comparative characterization and possible involvement in ascorbate metabolism.

Dabrowski K, Lackner R, Doblander C

Institute of Zoology, University of Innsbruck, Austria.

Comp Biochem Physiol B. 1993 Apr;104(4):717-22. doi: 10.1016/0305-0491(93)90203-h.

Abstract

The new assay conditions were determined for crude and purified enzyme ascorbate-2-sulfate sulfohydrolase from liver tissues of two fish species and bovine. 2. The major departure from the existing indirect method, based on reduction of 2,6-dichlorophenolindophenol (DCIP) by released ascorbic acid and change from pink-blue to a colorless molecule, takes into account the shift of maximum absorbance of DCIP from 516 nm at pH 5.14 to 600 nm at pH 6.5. 3. The direct method is based on colorimetric assay of liberated ascorbic acid including correction for interfering substances. The optimum pH for both fish ascorbate sulfatases was 5.5. 4. The Km for bovine ascorbate sulfatase was confirmed to be approximately 7 mM at 37 degrees C. 5. Partly purified ascorbate-sulfate sulfohydrolase has a Km value in rainbow trout of 0.4 mM and it changes very little in the range of water temperatures characteristic for this stenothermic fish species. 6. In eurythermic chub, the Km values increased from 1.2 to 4.3 mM with rising temperatures.

摘要

确定了两种鱼类和牛肝脏组织中粗制和纯化的酶抗坏血酸-2-硫酸硫酸水解酶的新测定条件。2. 与现有的间接方法相比，主要区别在于，现有间接方法基于释放的抗坏血酸还原2,6-二氯酚靛酚（DCIP），使其从粉红蓝色变为无色分子，而新方法考虑到DCIP在pH 5.14时最大吸光度在516 nm，在pH 6.5时变为600 nm。3. 直接方法基于对释放的抗坏血酸进行比色测定，包括对干扰物质的校正。两种鱼类抗坏血酸硫酸酯酶的最适pH均为5.5。4. 已证实牛抗坏血酸硫酸酯酶在37℃时的Km约为7 mM。5. 部分纯化的抗坏血酸-硫酸硫酸水解酶在虹鳟中的Km值为0.4 mM，并且在这种狭温性鱼类特有的水温范围内变化很小。6. 在广温性的鲢鱼中，Km值随温度升高从1.2 mM增加到4.3 mM。