Kraus R J, Prohaska J R, Ganther H E
Biochim Biophys Acta. 1980 Sep 9;615(1):19-26. doi: 10.1016/0005-2744(80)90004-2.
[75Se]Glutathione peroxidase (glutathione:hydrogen-peroxide oxidoreductase, EC 1.11.1.9) containing 4 mol selenium per mol was isolated in 33% yield using 10% ethanol to stabilize the purified enzyme. When reduced with GSH and rapidly separated from GSH by gel filtration chromatography, GSH peroxidase was eluted in a labile oxidized (iodoacetate-insensitive) form which was stable at 4 degrees C but unstable at 25 degrees C (form A). When GSH-reduced enzyme was allowed to oxidize in the course of dialysis a more stable oxidized form was obtained (form C) which was rapidly inactivated by cyanide. Using [35S]GSH, form C was shown to contain tightly bound glutathione in approx. equimolar ratio with selenium. The cyanide sensitivity of GSH peroxidase is therefore correlated with the presence of a glutathione moiety in the enzyme. The isolation of GSH peroxidase containing bound glutathione suggests that intermediates containing glutathione bound to selenium may be formed during the catalytic cycle.
以33%的产率分离得到了每摩尔含有4摩尔硒的[75Se]谷胱甘肽过氧化物酶（谷胱甘肽：过氧化氢氧化还原酶，EC 1.11.1.9），使用10%乙醇来稳定纯化后的酶。用谷胱甘肽（GSH）还原后，通过凝胶过滤色谱法迅速将其与GSH分离，谷胱甘肽过氧化物酶以不稳定的氧化（对碘乙酸不敏感）形式洗脱出来，该形式在4℃稳定，但在25℃不稳定（A形式）。当谷胱甘肽还原的酶在透析过程中被氧化时，会得到一种更稳定的氧化形式（C形式），它会被氰化物迅速灭活。使用[35S]GSH表明，C形式大约含有与硒等摩尔比的紧密结合的谷胱甘肽。因此，谷胱甘肽过氧化物酶对氰化物的敏感性与酶中谷胱甘肽部分的存在相关。含有结合谷胱甘肽的谷胱甘肽过氧化物酶的分离表明，在催化循环过程中可能会形成含有与硒结合的谷胱甘肽的中间体。
