Department of Chemical Engineering, Pohang University of Science and Technology, San 31, Hyoja Dong, Pohang, 790-784, Korea.
Extremophiles. 2012 May;16(3):447-54. doi: 10.1007/s00792-012-0444-1. Epub 2012 Apr 6.
Thermoplasma acidophilum utilizes L-rhamnose as a sole carbon source. To determine the metabolic pathway of L-rhamnose in Archaea, we identified and characterized L-rhamnose dehydrogenase (RhaD) in T. acidophilum. Ta0747P gene, which encodes the putative T. acidophilum RhaD (Ta_RhaD) enzyme belonging to the short-chain dehydrogenase/reductase family, was expressed in E. coli as an active enzyme catalyzing the oxidation of L-rhamnose to L-rhamnono-1,4-lactone. Analysis of catalytic properties revealed that Ta_RhaD oxidized L-rhamnose, L-lyxose, and L-mannose using only NADP(+) as a cofactor, which is different from NAD(+)/NADP(+)-specific bacterial RhaDs and NAD(+)-specific eukaryal RhaDs. Ta_RhaD showed the highest activity toward L-rhamnose at 60 °C and pH 7. The K (m) and k (cat) values were 0.46 mM, 1,341.3 min(-1) for L-rhamnose and 0.1 mM, 1,027.2 min(-1) for NADP(+), respectively. Phylogenetic analysis indicated that branched lineages of archaeal RhaD are quite distinct from those of Bacteria and Eukarya. This is the first report on the identification and characterization of NADP(+)-specific RhaD.
嗜热酸菌利用 L-鼠李糖作为唯一的碳源。为了确定 L-鼠李糖在古菌中的代谢途径，我们鉴定并表征了嗜热嗜酸菌中的 L-鼠李糖脱氢酶（RhaD）。Ta0747P 基因编码假定的嗜热嗜酸菌 RhaD（Ta_RhaD）酶，属于短链脱氢酶/还原酶家族，在大肠杆菌中表达为一种活性酶，可催化 L-鼠李糖氧化为 L-鼠李糖-1,4-内酯。催化特性分析表明，Ta_RhaD 仅使用 NADP(+)作为辅因子氧化 L-鼠李糖、L-岩藻糖和 L-甘露糖，这与 NAD(+)/NADP(+)-特异性细菌 RhaDs 和 NAD(+)-特异性真核 RhaDs 不同。Ta_RhaD 在 60°C 和 pH 7 时对 L-鼠李糖表现出最高的活性。K(m)和 k(cat)值分别为 0.46 mM 和 1,341.3 min(-1)，对于 L-鼠李糖和 0.1 mM 和 1,027.2 min(-1)，对于 NADP(+)。系统发育分析表明，古菌 RhaD 的分支谱系与细菌和真核生物的 RhaD 截然不同。这是首次报道 NADP(+)-特异性 RhaD 的鉴定和表征。
