Core Viewpoint - The research team from Shandong University has developed a novel non-canonical reductive TCA (Nc-rTCA) pathway for high-yield succinic acid biosynthesis in Yarrowia lipolytica, addressing the limitations of the traditional reductive TCA pathway [2][5]. Group 1: Research Background - C4 dicarboxylic acids, particularly succinic acid, are valuable compounds used in pharmaceuticals, chemicals, and food industries. Succinic acid serves as a platform compound for synthesizing various high-value chemicals [3]. - Yarrowia lipolytica is identified as a promising host for succinic acid biosynthesis due to its excellent acid tolerance and compatibility with low-cost raw materials [3]. Group 2: Challenges and Solutions - The traditional reductive TCA (rTCA) pathway faces challenges due to insufficient NADH levels in yeast, leading to lower than theoretical conversion rates for succinic acid production [3][5]. - The research team previously constructed a high-yield succinic acid-producing strain using alternative pathways but noted that conversion rates still had room for improvement [5]. Group 3: Novel Pathway Development - The team proposed the Nc-rTCA pathway to circumvent the redox imbalance associated with the traditional rTCA pathway. This new pathway utilizes endogenous aspartate transaminase (YlAAT) and heterologous aspartate ammonia-lyase (AAL) to convert oxaloacetate into fumarate, bypassing the NADH-dependent malate dehydrogenase [5]. - The introduction of NADPH-dependent glutamate dehydrogenase (YlGDH) facilitates the synthesis and utilization cycle of NH4, significantly improving the conversion rate of succinic acid [5]. Group 4: Experimental Results - The engineered Yarrowia lipolytica strain Ncr12 achieved a succinic acid yield of 98.16 g/L and a conversion rate of 0.91 g/g glucose in a 5-L fermentation tank, marking the highest conversion rate for succinic acid production from glucose in yeast [6]. - Using non-food low-cost raw materials such as crude glycerol and lignocellulose, the strain produced succinic acid yields of 117.74 g/L and 74.34 g/L, with conversion rates of 0.74 g/g glycerol and 0.98 g/g glucose, respectively, also representing the highest conversion rates for these carbon sources in yeast [6]. Group 5: Broader Implications - The Nc-rTCA pathway is theoretically universal and scalable, providing a new route for succinic acid production and a platform for the industrial production of other bio-based C4 dicarboxylic acids [6].