Yakunin A F, Hallenbeck P C
Département de Microbiologie et Immunologie, Université de Montreal, Québec, Canada.
J Bacteriol. 1997 Mar;179(5):1460-8. doi: 10.1128/jb.179.5.1460-1468.1997.
The synthesis of pyruvate carboxylase (PC) was studied by using quantitative immunoblot analysis with an antibody raised against PC purified from Rhodobacter capsulatus and was found to vary 20-fold depending on the growth conditions. The PC content was high in cells grown on pyruvate or on carbon substrates metabolized via pyruvate (lactate, D-malate, glucose, or fructose) and low in cells grown on tricarboxylic acid (TCA) cycle intermediates or substrates metabolized without intermediate formation of pyruvate (acetate or glutamate). Under dark aerobic growth conditions with lactate as a carbon source, the PC content was approximately twofold higher than that found under light anaerobic growth conditions. The results of incubation experiments demonstrate that PC synthesis is induced by pyruvate and repressed by TCA cycle intermediates, with negative control dominating over positive control. The content of PC in R. capsulatus cells was also directly related to the growth rate in continuous cultures. The analysis of intracellular levels of pyruvate and TCA cycle intermediates in cells grown under different conditions demonstrated that the content of PC is directly proportional to the ratio between pyruvate and C4 dicarboxylates. These results suggest that the regulation of PC synthesis by oxygen and its direct correlation with growth rate may reflect effects on the balance of intracellular pyruvate and C4 dicarboxylates. Thus, this important enzyme is potentially regulated both allosterically and at the level of synthesis.
利用针对从荚膜红细菌纯化的丙酮酸羧化酶（PC）产生的抗体，通过定量免疫印迹分析研究了丙酮酸羧化酶的合成，发现其合成量根据生长条件的不同而变化20倍。在以丙酮酸或通过丙酮酸代谢的碳底物（乳酸、D-苹果酸、葡萄糖或果糖）上生长的细胞中，PC含量较高；而在以三羧酸（TCA）循环中间体或不经过丙酮酸中间形成过程代谢的底物（乙酸或谷氨酸）上生长的细胞中，PC含量较低。在以乳酸作为碳源的黑暗需氧生长条件下，PC含量比在光照厌氧生长条件下大约高两倍。孵育实验结果表明，PC的合成由丙酮酸诱导，被TCA循环中间体抑制，负调控占主导地位。荚膜红细菌细胞中PC的含量也与连续培养中的生长速率直接相关。对在不同条件下生长的细胞内丙酮酸和TCA循环中间体水平的分析表明，PC的含量与丙酮酸和C4二羧酸之间的比例成正比。这些结果表明，氧气对PC合成的调节及其与生长速率的直接相关性可能反映了对细胞内丙酮酸和C4二羧酸平衡的影响。因此，这种重要的酶可能在别构水平和合成水平上都受到调控。
