The Construction of Inositol Oxidase Engineering Bacteria and optimization of Biotransformation System for Producing Glucuronic Acid
D-glucuronic acid is widely used in medicine,health food and cosmetics due to its good detoxification properties.With the rapid development of synthetic biology,it has become a trend to replace chemical synthesis with biological fermentation to produce glucuronic acid by engineering bacteria.This study aims to construct a biotransformation system of glucuronic acid,and to optimize its process.The results show that:the optimal induction conditions were when OD600 reached 0.5,0.2 mmol/L IPTG wa added in the medium,and the induction was performed at 26 ℃ for 8 hours,under these conditions,the recombinant strain BL21/pETDuet-miox,which could achieve high expression of inosiol oxidase(MIOX).The inositol oxidase vector plasmid pETDuet-miox was transformed into different expression hosts,and the best expression host was Escherichia coli Rosetta(DE3)according to the bioconversion yield.Using inositol oxidase as catalyst to catalyze the production of glucuronic acid from inositol,the best catalytic form is freeze-broken bacteria.The best catalyzing conditions were as follows:in the MOPS buffer system with pH value 7.5,the catalyzing temperature was 30 ℃,the amount of bacteria added was 100 g/L wet bacteria,and the substrate concentration was 2 g/L.In these circumstances the inositol conversion rate was 80.3％and the glucuronic acid production was 1.606 g/L.This study provides a new method for biosynthesis of glucuronic acid and lays a foundation for low-cost industrial production of glucuronic acid.
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