恶臭假单胞菌NA-1菌体培养转化和静息细胞转化联合工艺生产6-羟基烟酸研究

现有微生物羟基化烟酸采用的是静息细胞转化工艺。但研究揭示,恶臭假单胞菌NA-1(Pseudomonas putidaNA-1)在培养过程中不降解发酵液中由诱导剂烟酸转化形成的6-羟基烟酸,这是由于烟酸的存在抑制了羟基烟酸降解酶的作用,而不是因为细胞停止生长不利用羟基烟酸的缘故。因而尝试利用菌体诱导培养过程进行烟酸转化生产,建立了一种新的生产工艺,即菌体培养转化和静息细胞转化联合工艺。该工艺在恶臭假单胞菌NA-1培养过程中持续补充烟酸以维持1%(W/V)浓度,使烟酸被生长细胞转化为羟基化烟酸并在发酵液中线性积累,而不被进一步降解;培养转化结束后,发酵液中的静息细胞依然拥有很高的羟基化酶活力,能够再次用于转化反应。该联合转化工艺与传统的静息细胞转化工艺相比,不仅节约了诱导剂烟酸,而且6-羟基烟酸的产量提高了65%。

Combination of growing culture transformation and resting cells transformation of Pseudomonas putida NA-1 for production of 6-hydroxynicotinic acid

The current technology for microbial hydroxylation of nicotinic acid is a procedure of resting cells transformation.Here it was reported that non-degradation of 6-hydroxynicotinic acid during hydroxylation of nicotinic acid by resting cells was due to existing of nicotinic acid in the transformation mixture,not the death of cells.Therefore it was attempted that the induced cultivation of Pseudomonas putida NA-1 for hydroxylation activity by nicotinic acid was used for accumulation of 6-hydroxynicotinic acid to establish a new technology,a combined technology of growing culture transformation and resting cells transformation.In the combination technology,when 1% of nicotinic acid was kept in cultivation medium by supplement of nicotinic acid at intervals during cultivation of Pseudomonas putida NA-1,the growing culture could consistently hydroxylate nicotinic acid without degradation of its product which was accumulated linearly in the cultivation medium with progress of cultivation;the resting cells obtained from cultivation broth after growing culture transformation still processed a very high hydroxylation activity and could be used to next resting transformation of nicotinic acid.By using of the combined technology,the inducer was saved and the production yield of 6-hydroxynicotinic acid increased above 65%.