醋酸梭菌在异养和自养条件下的乙酸生成与代谢特性

Acetate production and metabolic characteristics of Clostridium aceticum under heterotrophic and autotrophic conditions

摘要:

【背景】产乙酸菌可以利用一碳气体发酵生产乙酸等高值化学品，是实现碳资源回收利用和绿色生物制造的重要途径之一。绝大部分产乙酸菌还可以利用有机质进行代谢，但二者具有不同的能量代谢模式。【目的】探究一种典型产乙酸菌醋酸梭菌(Clostridium aceticum)在异养/自养条件下的产酸特性，揭示不同培养条件对其代谢模式的影响机理。【方法】分别在以葡萄糖和合成气(H₂:CO₂为6:4)为底物的条件下考察醋酸梭菌产酸和代谢的情况，基于实验结果分析产物分布及乙酸生产比滴度，进一步利用关键酶活性指标揭示异养/自养代谢的差异性机理。【结果】醋酸梭菌在10 g/L葡萄糖存在时进行分步降解，先分解葡萄糖产生乳酸，后续生成6.3 g/L乙醇、13.2 g/L丁酸和5.6 g/L乙酸，而在合成气为底物时只检测到1.96 g/L乙酸。产物分布结果表明自养条件下的乙酸生产比滴度和酸醇比分别最高可达28.6 g/g和17.57，远高于异养的8.23 g/g和1.53。酶代谢指标表明，异养条件下的NADH、ATP产生量及丙酮酸脱氢酶活性均表现出明显优势，但乙酸激酶活性却恰好相反，这揭示出异养条件下丙酮酸代谢的中间产物乙酰辅酶A大部分流向细胞合成和高级产物(乙醇和丁酸)，反而在自养条件下流向乙酸生成。【结论】醋酸梭菌在自养条件下可以实现更专注的乙酸生成，为合成气的生物资源化利用及产乙酸菌的代谢通路调控提供一定的参考价值。

Abstract:

[Background] Acetic acid-producing bacteria can utilize one-carbon gas to produce high-value chemicals such as acetic acid by fermentation, which is an important way for carbon resource recycling and green biomanufacturing. In addition, most of the acetic acid-producing bacteria can utilize organic matter for metabolism. However, the two ways have different energy metabolism patterns. [Objective] To investigate the acid production characteristics of a typical acetic acid-producing bacterium, Clostridium aceticum, under heterotrophic/autotrophic conditions, and to reveal the mechanism of different culture conditions in affecting the metabolic patterns of this bacterium. [Methods] The acid production and metabolism of C.aceticum cultured with glucose and syngas (H₂ and CO₂ mixed in the ratio of 6:4), respectively, were investigated. The distribution of the products and the specific titer of acetic acid production were analyzed on the basis of the results of the experiments. The key enzyme activity indexes were employed to reveal the mechanism of the differences in the heterotrophic and autotrophic metabolism patterns. [Results] C.aceticum performed stepwise degradation in the presence of 10 g/L glucose. It firstly decomposed glucose to produce lactic acid, and subsequently produced 6.3 g/L ethanol, 13.2 g/L butyric acid, and 5.6 g/L acetic acid. However, only 1.96 g/L acetic acid was detected in the case of syngas as the substrate. The results of product distribution showed that the specific titer of acetic acid production and acid/alcohol ratio under autotrophic conditions reached up to 28.6 g/g and 17.57, respectively, which were much higher than those (8.23 g/g and 1.53) under heterotrophic conditions. The NADH and ATP production as well as pyruvate dehydrogenase activity were higher under heterotrophic conditions, while the acetate kinase activity showed an opposite trend. This result revealed that most of acetyl coenzyme A as an intermediate of pyruvate metabolism, flowed to cell synthesis and end products (ethanol and butyric acid) under heterotrophic conditions, while it flowed to acetate production under autotrophic conditions. [Conclusion] C.aceticum could achieve higher acetic acid production under autotrophic conditions, which provides a reference value for the utilization of syngas by microorganisms and the regulation of metabolic pathway of acetic acid-producing bacteria.