Disruption of the operon encoding Ehb hydrogenase limits anabolic CO2 assimilation in the archaeon Methanococcus maripaludis
Disruption of the operon encoding Ehb hydrogenase limits anabolic CO2 assimilation in the archaeon Methanococcus maripaludis
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DOI：
10.1128/jb.188.4.1373-1380.2006
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发表时间：
2006-02-01
影响因子：
3.2
通讯作者：
Whitman, WB
中科院分区：
文献类型：
--
作者：
Porat, I;Kim, W;Whitman, WB
甲烷球菌 maripaludis 是一种嗜温古菌，可以利用氢气或甲酸盐作为能源，将二氧化碳还原为甲烷。它含有两种膜结合的节能氢化酶，Eha 和 Ebb。为了确定 Ebb 的作用，构建了 ehb 操纵子的缺失以产生突变体，即菌株 S40。 S40 在基本培养基中的生长受到严重损害。醋酸盐和酵母提取物都是必需的，才能将生长恢复到接近野生型的水平，这表明 Ebb 参与了碳同化的多个步骤。然而，在细胞提取物或膜纯化级分中，野生型和突变体之间的总氢化酶比活性没有差异。在 S40 中，以丙酮酸作为电子供体的静息细胞的产甲烷作用也减少了 30%，表明丙酮酸氧化存在缺陷。 CO脱氢酶/乙酰辅酶A (CoA)合酶和丙酮酸氧化还原酶在突变体中具有更高的比活性，并且编码这些酶的基因以及形成AMP的乙酰辅酶A合成酶的表达水平增加。这些观察结果支持 Ebb 在甲烷球菌的合成代谢 CO2 同化中的作用。
Methanococcus maripaludis is a mesophilic archaeon that reduces CO2 to methane with H, or formate as an energy source. It contains two membrane-bound energy-conserving hydrogenases, Eha and Ebb. To determine the role of Ebb, a deletion in the ehb operon was constructed to yield the mutant, strain S40. Growth of S40 was severely impaired in minimal medium. Both acetate and yeast extract were necessan, to restore growth to nearly wild-type levels, suggesting that Ebb was involved in multiple steps in carbon assimilation. However, no differences in the total hydrogenase specific activities were found between the wild type and mutant in either cell extracts or membrane-purified fractions. Methanogenesis by resting cells with pyruvate as the electron donor was also reduced by 30% in S40, suggesting a defect in pyruvate oxidation. CO dehydrogenase/acetyl coenzyme A (CoA) synthase and pyruvate oxidoreductase had higher specific activities in the mutant, and genes encoding these enzymes, as well as AMP-forming acetyl-CoA synthetase, were expressed at increased levels. These observations support a role for Ebb in anabolic CO2 assimilation in methanococci.