Acetate Activation in Methanosaeta thermophila: Characterization of the Key Enzymes Pyrophosphatase and Acetyl-CoA Synthetase
Acetate Activation in Methanosaeta thermophila: Characterization of the Key Enzymes Pyrophosphatase and Acetyl-CoA Synthetase
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DOI：
10.1155/2012/315153
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发表时间：
2012-01-01
影响因子：
2.4
通讯作者：
Deppenmeier, Uwe
中科院分区：
文献类型：
--
作者：
Berger, Stefanie;Welte, Cornelia;Deppenmeier, Uwe
嗜热产甲烷菌嗜热甲烷菌以乙酸盐为唯一底物进行产甲烷。有人认为，将醋酸盐输入产甲烷途径所需的醋酸盐活化反应需要两个ATP的水解，而甲烷八叠球菌中的醋酸盐活化反应则需要两个ATP的水解酶。已知只需要一个三磷酸腺苷。由于这些生物生活在维持生命的热力学极限，山上的醋酸盐活化反应。嗜热菌似乎太贵了，因此被重新评估。研究发现，在可能的醋酸酯激活酶中，有一个编码形成AMP的乙酰-辅酶A合成酶的基因高度表达。纯化了相应的酶，并对其进行了详细的鉴定。它催化依赖于ATP的乙酰辅酶A、AMP和焦磷酸(PPI)的形成，而PPI仅适度抑制。PPI的分解是由一种可溶性焦磷酸酶完成的。并对该酶进行了纯化和鉴定。焦磷酸酶能降解醋酸酯活化反应中产生的大部分PPI(K-M=0.27+/-0.05 mm)。核苷酸和PPI对酶活性无抑制作用。然而，不能排除其他依赖PPI的酶利用剩余的PPI并有助于细胞的能量平衡。
The thermophilic methanogen Methanosaeta thermophila uses acetate as sole substrate for methanogenesis. It was proposed that the acetate activation reaction that is needed to feed acetate into the methanogenic pathway requires the hydrolysis of two ATP, whereas the acetate activation reaction in Methanosarcina sp. is known to require only one ATP. As these organisms live at the thermodynamic limit that sustains life, the acetate activation reaction in Mt. thermophila seems too costly and was thus reevaluated. It was found that of the putative acetate activation enzymes one gene encoding an AMP-forming acetyl-CoA synthetase was highly expressed. The corresponding enzyme was purified and characterized in detail. It catalyzed the ATP-dependent formation of acetylCoA, AMP, and pyrophosphate (PPi) and was only moderately inhibited by PPi. The breakdown of PPi was performed by a soluble pyrophosphatase. This enzyme was also purified and characterized. The pyrophosphatase hydrolyzed the major part of PPi (K-M = 0.27 +/- 0.05 mM) that was produced in the acetate activation reaction. Activity was not inhibited by nucleotides or PPi. However, it cannot be excluded that other PPi-dependent enzymes take advantage of the remaining PPi and contribute to the energy balance of the cell.