Metabolic evolution and super(13)C flux analysis of a succinate dehydrogenase deficient strain of Yarrowia lipolytica

• Published in: Biotechnology and bioengineering 2016-11, Vol.113 (11), p.2425-2432
• Main Authors: Yuzbashev, Tigran V, Bondarenko, Pavel Yu, Sobolevskaya, Tatiana I, Yuzbasheva, Evgeniya Yu, Laptev, Ivan A, Kachala, Vadim V, Fedorov, Alexander S, Vybornaya, Tatiana V, Larina, Anna S, Sineoky, Sergey P
• Format: Article
• Language: English
• Subjects: Bioengineering; Derivatives; Evolution; Glucose; Glycerols; Pathways; Succinic acid; Yarrowia lipolytica
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.26007

Bio-based succinic acid production can redirect industrial chemistry processes from using limited hydrocarbons to renewable carbohydrates. A fermentation process that does not require pH-titrating agents will be advantageous to the industry. Previously, a Yarrowia lipolytica strain that was defective for succinate dehydrogenase was constructed and was found to accumulate up to 17.5gL super(-1) of succinic acid when grown on glycerol without buffering. Here, a derivative mutant was isolated that produced 40.5gL super(-1) of succinic acid in 36h with a yield of 0.32gg super(-1) glycerol. A combination approach of induced mutagenesis and metabolic evolution allowed isolation of another derivative that could utilize glucose efficiently and accumulated 50.2gL super(-1) succinic acid in 54h with a yield of 0.43gg super(-1). The parent strain of these isolated mutants was used for [1,6- super(13)C sub(2)]glucose assimilation analysis. At least 35% glucose was estimated to be utilized through the pentose phosphate pathway, while greater than or equal to 84% succinic acid was formed through the oxidative branch of the tricarboxylic acid cycle. Biotechnol. Bioeng. 2016; 113: 2425-2432. This study demonstrated that the combination of induced mutagenesis and metabolic evolution strongly enhanced succinic acid production by Yarrowia lipolytica deficient for succinate dehydrogenase. The strain grew well without pH control, and accumulated up to 50.2gL super(-1) of succinic acid in 54h with a yield 0.43gg super(-1) glucose. The super(13)C-tracing experiment demonstrated that at least 35% of glucose was utilized through the PP pathway, while greater than or equal to 84% succinic acid formed from the oxidative branch of the TCA cycle.