MOLECULAR PHYSIOLOGY OF SUCCINIC ACID ACID BASED FERMENTATIONS IN ANAEROBES

丁二酸，第一大批量商品化学品， 可以通过发酵植物生物质底物制成，目前是 从石化原料中生产。琥珀酸是一种 柠檬酸循环的中间体，以及许多厌氧细菌 形成琥珀酸作为一种发酵产物。细菌 琥珀酸产厌氧螺杆菌可生产35-50g/L 琥珀酸，而放线杆菌属。产量130Z 65-80g/L。 磷酸烯醇式丙酮酸(PEP)羧酸激酶是关键的、受调节的酶 将PEP和二氧化碳缩合形成草酰乙酸酯，在导致 琥珀酸。成功地克隆和表达了A. 琥珀酸类在大肠杆菌中产生PEP羧激酶。这60.9 kDa 含有N-末端组氨酸标签的单体重组酶 纯化并用于开发基于MS的蛋白质 测序法。目前我们已经对放线杆菌进行了纯化。 130Z PEP羧基酶。TMPP-Ac电荷衍生化与多肽 MALDI-PSD-MS测序用于辅助克隆 相应的基因。

Succinic acid, the first large quantity commodity chemical, which can be made by fermentation of plant biomass substrates, is presently produced from petrochemical feedstocks. Succinic acid is an intermediate of the citric acid cycle, and many anaerobic bacteria form succinate as a fermentation product. The bacterium Anaerobiospirillum succiniciproducens is able to produce 35-50 g/l of succinate, but Actinobacillus sp. produces 130Z 65-80g/l. Phosphoenolpyruvate (PEP) carboxykinase is the key, regulated enzyme condensing PEP and CO2 to form oxaloacetate, in the pathway leading to succinate. We have successfully cloned and expressed A. succiniciproducens PEP carboxykinase in E. coli. This 60.9 kDa monomeric recombinant enzyme, containing an N-terminal His tag, was purified and used for the development of an MS-based protein sequencing method. At present we have purified the Actinobacillus sp 130Z PEP carboxykinase. TMPP-Ac charge derivatization and peptide sequencing by MALDI-PSD-MS is being used to assist in cloning the corresponding gene.