Acetylacetone-cleaving enzyme Dke1:: a novel C-C-bond-cleaving enzyme from Acinetobacter johnsonii
Acetylacetone-cleaving enzyme Dke1:: a novel C-C-bond-cleaving enzyme from Acinetobacter johnsonii
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DOI：
10.1042/bj20021047
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发表时间：
2003-02-01
影响因子：
4.1
通讯作者：
Steiner, W
中科院分区：
文献类型：
--
作者：
Straganz, GD;Glieder, A;Steiner, W
乙酰丙酮的毒性已在多项研究中得到证实。然而，关于其解毒或矿化的代谢途径知之甚少。这里介绍的数据首次描述了乙酰丙酮的微生物降解和启动代谢途径的新型酶的表征。从以乙酰丙酮作为唯一碳源生长的约氏不动杆菌菌株中，将诱导型乙酰丙酮裂解酶纯化至均一。相应的基因，编码一个153个氨基酸的序列，不显示任何显着的关系，其他已知的蛋白质序列，被克隆和过表达在大肠杆菌中，并给出了高产率的活性酶。这种酶将乙酰丙酮分解成等摩尔量的丙酮醛和乙酸盐，消耗一当量的分子氧。不需要外源辅因子，但Fell与活性蛋白结合，并且对其催化活性至关重要。该酶对乙酰丙酮具有高亲和力，K-m为9.1 μ M，k（cat）为8.5 s（-1）。乙酰丙酮降解的代谢途径和推定的关系，这种新的酶，以前描述的双加氧酶进行了讨论。
The toxicity of acetylacetone has been demonstrated in various studies. Little is known, however, about metabolic pathways for its detoxification or mineralization. Data presented here describe for the first time the microbial degradation of acetylacetone and the characterization of a novel enzyme that initiates the metabolic pathway. From an Acinetobacter johnsonii strain that grew with acetylacetone as the sole carbon source, an inducible acetylacetone-cleaving enzyme was purified to homogeneity. The corresponding gene, coding for a 153 amino acid sequence that does not show any significant relationship to other known protein sequences, was cloned and overexpressed in Escherichia coli and gave high yields of active enzyme. The enzyme cleaves acetylacetone to equimolar amounts of methylglyoxal and acetate, consuming one equivalent of molecular oxygen. No exogenous cofactor is required, but Fell is bound to the active protein and essential for its catalytic activity. The enzyme has a high affinity for acetylacetone with a K-m of 9.1 muM and a k(cat) of 8.5 s(-1). A metabolic pathway for acetylacetone degradation and the putative relationship of this novel enzyme to previously described dioxygenases are discussed.