Cell factories and transcription factor engineering for bioproducts: the case of Candida glabrata for α-ketoglutarate production

α-Ketoglutarate, a key intermediate in the TCA cycle, is crucial for amino acid synthesis and nitrogen transport. However, microbial engineering for α-ketoglutarate production is hindered by the intrinsic inefficiency of the metabolic network. In this study, transcription factor engineering was performed for the reconstruction of the metabolic network to boost α-ketoglutarate biosynthesis in Candida glabrata. Transcription factors GCR2 and RTG1 were first reinstalled to kick-start the glycolytic pathway and the TCA cycle, respectively, and then optimized to redistribute carbon flux between the two pathways. In addition, pyruvate carriers, MPC1 and MPC2, were introduced to facilitate the transport of cytoplasmic pyruvate to mitochondria, thereby feeding it into the TCA cycle for α-ketoglutarate biosynthesis. Next, transcription factor HAP4 was utilized to rewire the electron transport chain for improving redox balance and reducing overflow metabolism, thereby channeling more carbon flux to α-ketoglutarate production. Finally, the engineered strain C. glabrata KGA17 was capable of producing 210.4 g/L α-ketoglutarate in a 5-L bioreactor. This approach showed significant promise for developing efficient microbial cell factories for high-value chemical production.