Metabolic Regulation On Beta-Oxidation Pathway In The Production Of 1, 11-Dicarboxylic Acid Through Biocatalysis

Long chainα,ω-dicarboxylic acids (DCA) are a series of important fine chemicals. Candida tropicalis can be used for oxidizing n-alkanes or fatty acids with carbon chain length from 11 to 18 to produce the corresponding DCA, in which excessiveβ-oxidation of products will decrease the productivity. Theβ-oxidation pathway in bioproduction of DCA13 by Candida tropicalis was investigated. The traditional application of metabolic engineering for reconstruction of the key enzymes was broken through, and new application was explored. To further increase the conversion of DCA13 with respect to alkane, the transportation system of acetyl-CoA, which linksβ-oxidation and TCA cycle together, was engineered, to regulate the metabolic fluxes inβ-oxidation. This method can be served as a new way for reconstructing the transportation system in eucaryote and industrial microbiological technologies. From the view of providing assistant energy source as a substitute ofβ-oxidation, fed-batch fermentation was studied.The key enzyme in the acetyl-CoA transportation system, carnitine acetyltransferase (CAT) was engineered. One copy of CAT gene was knocked out, and recombinant strain CZ-15 was constructed. Both the features of morphology and growth of CZ-15 were familiar with those of F10-1, the original strain. Compared with F10-1, DCA13 concentration and alkane conversion of CZ-15 were increased by 18.3 % and 12.0 % respectively, and the characteristics of CZ-15 could be inherited stably.Based on the metabolic network models of Candida tropicalis, the fermentation data were analyzed. The results indicated that, compared with F10-1, in CZ-15, the acetyl-CoA transported into TCA by CAT was 50% lower, and theβ-oxidation of DCA13 was also decreased, showing that the reduction of CAT activity could decrease the excessive consumption of DCA13.Sequentially, both of the two copies of CAT genes were knocked out, and the recombinant strain CKC-11 was obtained, in which no CAT activity was detected. CKC-11 could not grow on alkane and produce DCA13, which indicated that the total...