A detailed biochemical characterization of phosphopantothenate synthetase, a novel enzyme involved in coenzyme A biosynthesis in the Archaea
A detailed biochemical characterization of phosphopantothenate synthetase, a novel enzyme involved in coenzyme A biosynthesis in the Archaea
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DOI：
10.1007/s00792-012-0477-5
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发表时间：
2012-11-01
期刊：
影响因子：
2.9
通讯作者：
Atomi, Haruyuki
中科院分区：
文献类型：
--
作者：
Ishibashi, Takuya;Tomita, Hiroya;Atomi, Haruyuki
我们以前曾报道，大多数古细菌利用一种新的途径辅酶A的生物合成（CoA）。细菌/真核生物通常使用泛酸合成酶和泛酸激酶将泛酸转化为4 '-磷酸泛酸。然而，在极端嗜热古菌Thermococcus kodakarensis中，两种新的古菌特有的酶，泛解酸激酶和磷酸泛酸合成酶，负责这种转换。在这里，我们研究了古生菌磷酸泛解酸合成酶的酶学性质，该合成酶催化4-磷酸泛解酸和β-丙氨酸的ATP依赖性缩合。磷酸泛酸合成酶反应的活化能为82.3 kJ mol（-1）。在对核苷三磷酸的底物特异性方面，该酶显示出对ATP的严格偏好。在几种胺底物中，用β-丙氨酸检测到活性，但用γ-氨基丁酸、甘氨酸或天冬氨酸均未检测到活性。磷酸泛解酸合成酶反应遵循Michaelis-Menten动力学对β-丙氨酸，而底物抑制与4-磷酸泛解酸和ATP观察。未观察到CoA/乙酰CoA的反馈抑制和4 '-磷酸泛酸的产物抑制。相比之下，其他古细菌酶泛解酸激酶显示4-磷酸泛解酸以非竞争性方式抑制产物。根据我们的研究结果，我们讨论了古细菌辅酶A生物合成的调控。
We have previously reported that the majority of the archaea utilize a novel pathway for coenzyme A biosynthesis (CoA). Bacteria/eukaryotes commonly use pantothenate synthetase and pantothenate kinase to convert pantoate to 4'-phosphopantothenate. However, in the hyperthermophilic archaeon Thermococcus kodakarensis, two novel enzymes specific to the archaea, pantoate kinase and phosphopantothenate synthetase, are responsible for this conversion. Here, we examined the enzymatic properties of the archaeal phosphopantothenate synthetase, which catalyzes the ATP-dependent condensation of 4-phosphopantoate and beta-alanine. The activation energy of the phosphopantothenate synthetase reaction was 82.3 kJ mol(-1). In terms of substrate specificity toward nucleoside triphosphates, the enzyme displayed a strict preference for ATP. Among several amine substrates, activity was detected with beta-alanine, but not with gamma-aminobutyrate, glycine nor aspartate. The phosphopantothenate synthetase reaction followed Michaelis-Menten kinetics toward beta-alanine, whereas substrate inhibition was observed with 4-phosphopantoate and ATP. Feedback inhibition by CoA/acetyl-CoA and product inhibition by 4'-phosphopantothenate were not observed. By contrast, the other archaeal enzyme pantoate kinase displayed product inhibition by 4-phosphopantoate in a non-competitive manner. Based on our results, we discuss the regulation of CoA biosynthesis in the archaea.