Research On Bioconversion Mechanism Of 3-hydroxypropionic Acid And Its Biocatalysis Technology By Acetobacter Sp.

3-hydroxypropionic acid(3-HP) is one of the emerging industrial chemical intermediates with high economic value and wide application prospect, which has not been produced in industry by now. 3-HP synthesis method using microbial catalysis, which has a wide substrate range, simple process and high conversion rate, is a technically feasible way of 3-HP production. A screening strains, Acetobacter sp. was screened in our early research and could efficiently catalytic 1,3-propanediol(1,3-PDO) into 3-HP. This study is aimed at probing preliminarily into the reaction mechanism of 3-HP biosynthesis process from 1,3-PDO oxidation by Acetobacter sp.. Immobilized cells were used in the established biocatalytic system for 3-HP synthesis and 3-HP biosynthesis process was explored, coupling of 1,3-PDO microbial fermentation from initial substrate glycerol and 3-HP biocatalysis, in order to provide a new method for microbial 3-HP production.Dehydrogenase enzyme activities of different Acetobacter sp. cell components were tested for the orientation of corresponding enzymes in 3-HP biocatalytic process; Enzyme activity of crude enzyme extract was tested using different electron acceptors for the conformation that the coenzyme of 1,3-PDO oxidation dehydrogenase was Pyrroloquinoline quinone(PQQ); The addition of ubiquinone, respiratory chain inhibitor retontone in enzyme activity test and final electron acceptor-oxygen supply blocking experiment proved that 1,3-PDO dehydrogenase had reductase activity and the process of dehydrogenation oxidation coupled with the cytoplasmic membrane-bound respiratory chain by ubiquinone. Electron came from substrates and was transferred from alcohols/aldehydes dehydrogenase to ubiquinone and then terminal oxidase and finally to oxygen. 3-HP biosynthesis was an oxidation process of 1,3-PDO oxidation through respiratory chain dehydrogenation reaction with high efficiency. According to the substrate concentration depression effect analysis of 1,3-PDO catalytic reaction, reaction was inhibited when over 50 g·L-1 1,3-PDO was oxidized.Basing on the mechanical strength, mass transfer property, 3-HP catalytic performance and cyclic utilization of immobilized cells, 3-HP biosynthesis system was established for promoting the substrate tolerance of catalytic reaction. 40 g·L-1 sodium alginate was the basic embedding material and mixed with 60 g·L-1 polyvinyl alcohol as compound material, 2 mm immobilized cells capsules showed mechanical strength 197.82 g and the mass transfer parameter was 257.54 px. The maximum 3-HP titer 66.95 g·L-1, with a molar yield of 80.77%, was reached when 70 g·L-1 1,3-PDO was catalyzed by immobilized cells. Besides, 50 g·L-1 1,3-PDO was circularly catalyzed in the established catalytic system and the yield was retained upon 80.65% at the 5th circle after 300 hours. Basing on the establishment and analysis of mathematical mass transfer model, the bio-oxidation process of 1,3-PDO and 3-HP synthesis was principally affected by the internal mass transfer when the internal effectiveness factor ηi < 1 and Thiele modulus φ > 0.3.Original strains of Acetobacter sp. was domesticated and culture medium was optimized for the establishment of experimental 3-HP synthesis system, coupling processes of immobilized cells catalysis and microbial fermentation from glycerol. Immobilized cells catalysis was combined with microbial 1,3-PDO production of Klebsiella pneumoniae and the running result of coupling system showed that 27.66 g·L-1 3-HP was catalyzed with molar yield of 58.62%, which verified the feasibility of fermentation and catalysis coupling method. A new route of 3-HP biosynthesis was provided using Acetobacter sp..