Asanuma Narito, Yoshizawa Kimio, Kanada Kazuhiro, Hino Tsuneo
Department of Life Science, College of Agriculture, Meiji University, Higashimita, Kawasaki, Japan.
Curr Microbiol. 2009 May;58(5):416-20. doi: 10.1007/s00284-008-9325-3. Epub 2009 Feb 5.
Molecular properties and transcriptional control of phosphoenolpyruvate carboxykinase (PCK; EC 4.1.1.32) in Ruminococcus albus were examined. The putative 537-amino acid PCK polypeptide has a predicted mass of 59.4 kDa and an isoelectric point of 4.82. RT-PCR and Northern blot analyses of pck mRNA suggest that the transcript is monocistronic and that pck transcription is not affected by changes in sugar sources present in growth medium. PCK enzymatic activity requires either Mg(2+) or Mn(2+) and an optimal pH of 7.0. R. albus PCK phosphorylated ADP more readily than GDP. Apparent K ( m ) values of PCK for PEP and ADP were considerably lower than those for OAA and ATP, suggesting that the reaction from PEP to OAA is favored in R. albus. The enzyme properties of PCK in R. albus appear to be more similar to Selenomonas ruminantium PCK than to Ruminococcus flavefacience, although R. albus and R. flavefacience belong to the same genus. The specific activity of PCK, representing the amount of enzyme per cell, in R. albus was much lower than that in S. ruminantium. The amount of succinate produced in R. albus from one unit of cellobiose was also much lower than the sum of succinate and propionate produced in S. ruminantium. Based on these results, we propose enhancement of PCK activity by stimulating PCK transcription as a method to decrease R. albus H(2) production without suppressing growth.
对白色瘤胃球菌中磷酸烯醇丙酮酸羧激酶（PCK；EC 4.1.1.32）的分子特性和转录调控进行了研究。推测的537个氨基酸的PCK多肽预测分子量为59.4 kDa，等电点为4.82。对pck mRNA的RT-PCR和Northern印迹分析表明，该转录本是单顺反子的，并且pck转录不受生长培养基中糖源变化的影响。PCK酶活性需要Mg(2+)或Mn(2+)，最适pH为7.0。白色瘤胃球菌PCK磷酸化ADP比磷酸化GDP更容易。PCK对PEP和ADP的表观K (m)值明显低于对OAA和ATP的表观K (m)值，这表明在白色瘤胃球菌中从PEP到OAA的反应更有利。尽管白色瘤胃球菌和黄化瘤胃球菌属于同一属，但白色瘤胃球菌中PCK的酶特性似乎与反刍月形单胞菌PCK更相似，而与黄化瘤胃球菌的PCK不太相似。白色瘤胃球菌中代表每个细胞中酶量的PCK比活性远低于反刍月形单胞菌中的比活性。白色瘤胃球菌从一个单位纤维二糖产生的琥珀酸量也远低于反刍月形单胞菌产生的琥珀酸和丙酸的总量。基于这些结果，我们提出通过刺激PCK转录来增强PCK活性，作为一种在不抑制生长的情况下减少白色瘤胃球菌氢气产生的方法。
