Krebs Institute, Department of Molecular Biology and Biotechnology, University of Sheffield, Firth Court, Western Bank, P.O. Box 594, S10 2UH, Sheffield, UK.
Photosynth Res. 1994 Jul;41(1):75-88. doi: 10.1007/BF02184147.
Heliobacteria are a group of anoxygenic phototrophs that can grow photoheterotrophically in defined minimal media on only a limited range of organic substrates as carbon sources. In this study the mechanisms which operate to assimilate carbon and the routes employed for the biosynthesis of cellular intermediates were investigated in a newHeliobacterium strain, HY-3. This was achieved using two approaches (1) by measuring the activities of key enzymes in cell-free extracts and (2) by the use of(13)C nuclear magnetic resonance (NMR) spectroscopy to analyze in detail the labelling pattern of amino-acids of cells grown on [(13)C] pyruvate and [(13)C] acetate.Heliobacterium strain HY-3 was unable to grow autotrophically on CO2/H2 and neither (ATP)-citrate lyase nor ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBPcase) were detectable in cell-free extracts. The enzyme profile of pyruvate grown cells indicated the presence of a pyruvate:acceptor oxidoreductase at high specific activity which could convert pyruvate to acetyl-Coenzyme A. No pyridine nucleotide dependent pyruvate dehydrogenase complex activity was detected. Of the citric-acid cycle enzymes, malate dehydrogenase, fumarase, fumarate reductase and an NADP-specific isocitrate dehydrogenase were readily detectable but no aconitase or citrate synthase activity was found. However, the labelling pattern of glutamate in long-term 2-[(13)C] acetate incorporation experiments indicated that a mechanism exists for the conversion of carbon from acetyl-CoA into 2-oxoglutarate. A 2-oxoglutarate:acceptor oxidoreductase activity was present which was also assayable by isotope exchange, but no 2-oxoglutarate dehydrogenase complex activity could be detected. Heliobacteria appear to use a type of incomplete reductive carboxylic acid pathway for the conversion of pyruvate to 2-oxoglutarate but are unable to grow autotrophically using this metabolic route due to the absence of ATP-citrate lyase.
噬氢菌是一类不产氧的光合自养菌，它们可以在仅有限的几种有机底物作为碳源的限定最小培养基中异养生长。在这项研究中，我们研究了一种新的噬氢菌 HY-3 菌株中同化碳的机制和细胞内中间产物生物合成的途径。这是通过两种方法实现的：（1）测量无细胞提取物中关键酶的活性，（2）使用（13）C 核磁共振（NMR）光谱分析详细分析细胞在 [(13)C]丙酮酸和 [(13)C]乙酸上生长时氨基酸的标记模式。噬氢菌 HY-3 菌株不能在 CO2/H2 上自养生长，也不能检测到无细胞提取物中的（ATP）柠檬酸裂解酶或核酮糖 1,5-二磷酸羧化酶/加氧酶（RuBPcase）。丙酮酸生长细胞的酶谱表明存在高比活性的丙酮酸：受体氧化还原酶，可将丙酮酸转化为乙酰辅酶 A。未检测到吡啶核苷酸依赖的丙酮酸脱氢酶复合物活性。在柠檬酸循环酶中，苹果酸脱氢酶、延胡索酸酶、延胡索酸还原酶和 NADP 特异性异柠檬酸脱氢酶很容易检测到，但未发现柠檬酸合酶或 aconitase 活性。然而，在长期 2-[(13)C]乙酸掺入实验中谷氨酸的标记模式表明，存在一种将乙酰辅酶 A 中的碳转化为 2-氧戊二酸的机制。存在 2-氧戊二酸：受体氧化还原酶活性，也可以通过同位素交换进行测定，但未检测到 2-氧戊二酸脱氢酶复合物活性。噬氢菌似乎使用一种不完全的还原羧酸途径将丙酮酸转化为 2-氧戊二酸，但由于缺乏 ATP-柠檬酸裂解酶，它们无法通过这种代谢途径自养生长。
