Purification and properties of the NAD+-dependent (type D) and O2-dependent (type O) forms of rat liver xanthine dehydrogenase.
Purification and properties of the NAD+-dependent (type D) and O2-dependent (type O) forms of rat liver xanthine dehydrogenase.
复制标题
大鼠肝黄嘌呤脱氢酶的 NAD 依赖性（D 型）和 O2 依赖性（O 型）形式的纯化和特性。
DOI：
10.1016/0003-9861(76)90087-4
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发表时间：
1976
影响因子：
3.9
通讯作者：
K. Rajagopalan
中科院分区：
文献类型：
--
作者：
William R. Waud;K. Rajagopalan
大鼠肝脏的黄嘌呤氧化酶已被纯化为 NAD+ 依赖性脱氢酶（D 型）和 O2 依赖性氧化酶（O 型）。通过聚丙烯酰胺不连续凝胶电泳判断，纯化的 D 和 O 变体几乎是同质的，并且在十二烷基硫酸钠-尿素凝胶上无法区分。 D 型酶的吸收光谱与 O 型酶的吸收光谱没有区别，并且与其他来源的黄嘌呤氧化酶的光谱非常相似。 D 型和 O 型酶具有基本相同的辅因子组成。 D 型黄嘌呤的氧化受到 NAD+ 的刺激，并伴随着 NADH 的形成。 D 型能够利用 NADH 以及黄嘌呤作为各种受体的电子供体，而 O 型则不能氧化 NADH。亚砷酸盐、氰化物和甲醇完全消除D型酶对黄嘌呤的氧化作用，同时不同程度地影响NADH的活性。在这些方面，大鼠肝黄嘌呤脱氢酶与鸡肝黄嘌呤脱氢酶非常相似。然而，与禽类酶相比，纯化的大鼠肝酶作为脱氢酶不稳定并逐渐转化为氧化酶。这种转化伴随着需氧黄嘌呤→细胞色素活性的增加。大鼠肝脏提取物中的天然 D 型酶可以用针对纯化的 O 型制备的抗体沉淀。两种形式的黄嘌呤 Km 没有显着差异。
The xanthine-oxidizing enzyme of rat liver has been purified as an NAD+-dependent dehydrogenase (type D) and as the O2-dependent oxidase (type O). The purified D and O variants are nearly homogenous as judged by polyacrylamide discontinuous gel electrophoresis and are indistinguishable on sodium dodecyl sulfate-urea gels. The absorption spectrum of the type D enzyme is indistinguishable from that of the type O enzyme and closely resembles the spectra of xanthine-oxidizing enzymes from other sources. The types D and O enzymes have essentially the same cofactor composition. Oxidation of xanthine by type D is stimulated by NAD+with concomitant NADH formation. Type D is able to utilize NADH as well as xanthine as electron donor to various acceptors, in contrast to type O that is unable to oxidize NADH. Arsenite, cyanide and methanol completely abolish xanthine oxidation by the type D enzyme while affecting the activities with NADH to varying extents. In these respects rat liver xanthine dehydrogenase closely resembles chicken liver xanthine dehydrogenase. However, in contrast to the avian enzyme, the purified rat liver enzyme is unstable as a dehydrogenase and is gradually converted to an oxidase. This conversion is accompanied by an increase in the aerobic xanthine → cytochromecactivity. The native type D enzyme in rat liver extracts is precipitable with antibody prepared against purified type O. TheKmfor xanthine is not significantly different for the two forms.