Metabolic Process Optimization Analysis Of Glycerol Bioconversion Into1,3-propanediol

Biodiesel is an alternative energy source and the biodiesel production istremendously increased in recent years. While10%of crude glycerol is accompaniedby a by-product in the process of biodiesel production, this leads to excess glycerolmarket inevitably. Taking glycerol as a raw material to produce a high valuableproduction becomes an urgent demand in biodiesel industry. Much attention has beenpaid on the bioconversion of glycerol into1,3-propanediol (1,3-PD). In view of thehigh cost of the bioconversion of glycerol into1,3-propanediol, it has become ahotspot of research to improve productivity of1,3-propanediol by metabolicengineering means. In this paper, metabolic pathways of glycerol in Klebsiellapneumoniae were optimized and analyzed, the metabolic pathways and key enzymesof biomass and1,3-propanediol were predicted respectively. These results providedtheoretical guidance for genetically engineered strains.The metabolic network mode of K. pneumoniae was first established by Booleannetwork by the oxygen regulation information. By applying linear programming withmaximization of biomass as the objective function, the elementary mode analysis inconjunction with experimentally accumulation rates of external metabolites for K.pneumoniae at different phases of fermentation was used to optimize and analyze themetabolic network under micro-aerobic and anaerobic conditions. The optimummetabolic pathways at different phases of fermentation were obtained. The optimummetabolic pathways contributing biomass synthesis were found identical at differentphases of fermentation under micro-aerobic condition. The optimum metabolicpathways of delay period and stable period were found to the same under anaerobic condition. The different optimum metabolic pathways were obtained at logarithmperiod compared with at delay period and at stable period under anaerobic condition.In addition, it was found that the optimum metabolic pathways largely contributed to1,3-propanediol production. The oxidative phosphorylation pathway played animportant role for maintaining reducing equivalent and energy balance and synthesis ofbiomass and1,3-propanediol.Next, we discussed the correlations between reactions of the metabolic network ofK. pneumoniae and synthesis of1,3-propanediol and biomass under micro-aerobic andanaerobic conditions. It was showed that the glycolytic oxidation-reduction pathwaywas propitious to the synthesis of1,3-propanediol and biomass regardless ofmicro-aerobic and anaerobic conditions; Tricarboxylic acid cycle had a greaterinfluence on synthesis of biomass. In addition, compared to anaerobic condition, thepentose phosphate pathway had a greater influence on synthesis of biomass undermicro-aerobic condition.At last, the metabolic network of K. pneumoniae under aerobic condition wassimulated by applying CASOP (Computational Approach for Strain Optimizationaiming at high Productivity) algorithm with high productivity of1,3-propanediol as theobjective function. We finally selected five knockout candidates (pyruvateformate-lyase gene, formate dehydrogenase gene, fumarate reductase gene, aldehydedehydrogenase gene,2,3-butanediol dehydrogenase gene) and five overexpressioncandidates (isocitrate lyase gene, malate synthetase gene, glucose-6-phosphatedehydrogenase gene,6-phosphate glucose acid esterase gene,6-phosphate glucose aciddehydrogenase gene).