Purification of ornithine decarboxylase from kidneys of androgen-treated mice.
Purification of ornithine decarboxylase from kidneys of androgen-treated mice.
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从雄激素处理小鼠的肾脏中纯化鸟氨酸脱羧酶。
DOI：
10.1021/bi00257a023
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发表时间：
1982
期刊：
影响因子：
2.9
通讯作者：
Pegg,AE
中科院分区：
文献类型：
--
作者：
Seely,JE;Pösö,H;Pegg,AE
James E. Seely,* Hannu Poso，§和Anthony E. Pegg*摘要：从雄激素处理的小鼠肾脏中纯化出鸟氨酸脱羧酶。这种肾脏的酶含量比其他哺乳动物组织（如诱导的大鼠肝脏）高2个数量级，并且只需要10350倍的纯化。该酶在天然条件和变性条件下均具有单条带等电聚焦和聚丙烯酰胺凝胶电泳。这些条带对应于酶活性和鸟氨酸脱羧酶的迁移，该酶与特异性抑制剂-（二氟甲基）[5-14C]鸟氨酸反应。该酶的Mr约为100000，是Mr 53000亚基的二聚体。l -鸟氨酸的Km为75/iM，磷酸吡哆醛的Km为0.3^ m。该制剂的比活性为50^ mol的co2产生min'1 mg'1，并结合了化学计量量的不可逆抑制剂a-（二氟甲基）鸟氨酸（每个亚基一个分子）。除非添加0.02%的brij35，否则即使在2.5 mM二硫苏糖醇和40 juM磷酸吡哆醛存在下，纯化的酶也不稳定。在这种洗涤剂存在的情况下，酶可以在几乎没有活性损失的情况下储存。鸟氨酸脱羧酶（EC 4.1）引起了人们极大的兴趣。1.17)，原因有很多[Morris & Fillingame（1974）、Janne et al.（1978）、Canellakis et al.（1979）、McCann（1980）、Russell（1980）和Pegg & Williams-Ashman（1981）评述]。在哺乳动物细胞中，这种酶是腐胺的唯一来源，腐胺是多胺的前体。它的活性在各种营养刺激下迅速增加，并且在哺乳动物酶中似乎具有最快的蛋白质合成和降解速度。其活性也可能受到大分子抑制剂水平（Canellakis et al., 1978）和多种翻译后修饰（Mitchell, 1981; Russell, 1981; Atmar & Kuehn, 1981）的调节。这种酶是潜在有用的化疗药物的靶标
James E. Seely,* Hannu Poso, § and Anthony E. Pegg* abstract: Ornithine decarboxylase has been purified to homogeneity from kidneys of androgen-treated mice. Such kidneys have an enzyme content 2 orders of magnitude greater than thatof other mammalian tissues such as induced rat liver, and only a 10350-fold purification was needed for purification. The enzyme preparation gave a single band on isoelectric focusing and on polyacrylamide gel electrophoresis under native and denaturing conditions. These bands corresponded to the enzyme activity and to the migration of ornithine de-carboxylase labeled by reaction with a-(difluoromethyl)[5-14C] ornithine, a specific inhibitor. The enzyme has a Mr of about 100000 and is a dimer of subunit Mr 53 000. The Km for L-ornithine was 75/iM and for pyridoxal phosphate, 0.3^ M. The preparation had a specific activityof 50^ mol of C02 produced min'1 mg'1 and bound a stoichiometric amount of the irreversible inhibitor, a-(difluoromethyl) ornithine (one molecule per subunit). The purified enzyme was unstable even in the presence of 2.5 mM dithiothreitol and 40 juM pyridoxal phosphate unless 0.02% Brij 35 was added. In thepresence of this detergent, the enzyme could be stored with little loss of activity. ere hasbeen considerableinterest in the enzyme ornithine decarboxylase (EC 4.1. 1.17) for a number of reasons [reviewed by Morris & Fillingame (1974), Janne et al.(1978), Canellakis et al.(1979), McCann (1980), Russell (1980), and Pegg & Williams-Ashman (1981)]. In mammalian cells, this enzyme provides the only source of putrescine, a precursor of the polyamines. Its activity increases veryrapidly in response to a wide variety of trophic stimuli, and it appears to have the most rapid rate of protein synthesis and degradation among mammalian enzymes. Its activity may also be regulated by the level of a macromolecular inhibitor (Canellakis et al., 1978) and a variety of posttranslational modifications (Mitchell, 1981; Russell, 1981; Atmar & Kuehn, 1981). The enzyme is the target for potentially useful chemotherapeutic agents