A new activity for an old enzyme:: Escherichia coli bacterial alkaline phosphatase is a phosphite-dependent hydrogenase
A new activity for an old enzyme:: Escherichia coli bacterial alkaline phosphatase is a phosphite-dependent hydrogenase
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DOI：
10.1073/pnas.0400664101
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发表时间：
2004-05-25
影响因子：
11.1
通讯作者：
Metcalf, WW
中科院分区：
文献类型：
--
作者：
Yang, KC;Metcalf, WW
遗传分析表明，大肠杆菌具有两条独立的亚磷酸盐(PT)氧化成磷的途径。其中一条途径依赖于14个基因的PHN操纵子，它编码C-P裂解酶。另一条途径依赖于编码细菌碱性磷酸酶(BAP)的PhoA基因座。转座子突变研究强烈表明，BAP是唯一参与PhoA依赖途径的酶。这一结论得到了铂氧化酶的纯化和生化特性的支持，N末端蛋白测序证实该酶为BAP。高纯度的BAP催化铂氧化，比活性为62-242mU/mg，催化磷酸酯水解，比活性为41-61u/mg。令人惊讶的是，BAP催化铂氧化成磷酸盐和分子H-2。因此，BAP是一种独特的铂依赖的放氢氢酶。这种反应在P和H两种生物化学中都是史无前例的，很可能涉及氢化物从底物到水生质子的直接转移。
Genetic analysis indicates that Escherichia coli possesses two independent pathways for oxidation of phosphite (Pt) to phosphate. One pathway depends on the 14-gene phn operon, which encodes the enzyme C-P lyase. The other pathway depends on the phoA locus, which encodes bacterial alkaline phosphatase (BAP). Transposon mutagenesis studies strongly suggest that BAP is the only enzyme involved in the phoA-dependent pathway. This conclusion is supported by purification and biochemical characterization of the Pt-oxidizing enzyme, which was proven to be BAP by N terminus protein sequencing. Highly purified BAP catalyzed Pt oxidation with specific activities of 62-242 milliunits/mg and phosphate ester hydrolysis with specific activities of 41-61 units/ mg. Surprisingly, BAP catalyzes the oxidation of Pt to phosphate and molecular H-2. Thus, BAP is a unique Pt-dependent, H-2-evolving hydrogenase. This reaction is unprecedented in both P and H biochemistry, and it is likely to involve direct transfer of hydride from the substrate to water-derived protons.