Enhancing β-alanine production from glucose in genetically modified Corynebacterium glutamicum by metabolic pathway engineering

• Published in: Applied microbiology and biotechnology 2021-12, Vol.105 (24), p.9153-9166
• Main Authors: Wang, Jin-Yu, Rao, Zhi-Ming, Xu, Jian-Zhong, Zhang, Wei-Guo
• Format: Article
• Language: English
• Subjects: Accumulation; Alanine; Aspartate kinase; Aspartic acid; Batch culture; Biomedical and Life Sciences; Biotechnological Products and Process Engineering; Biotechnology; Carbon; Corynebacterium glutamicum; Efficiency; Enzymes; Fermentation; Genetic modification; Glucose; Kinases; L-Alanine; L-Lactate dehydrogenase; Lactate dehydrogenase; Lactic acid; Life Sciences; Lysine; Metabolic engineering; Metabolic pathways; Microbial Genetics and Genomics; Microbiology; Microorganisms; Optimization; Valine
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-021-11696-y

To directly produce β-alanine from glucose by microbial fermentation, a recombinant Corynebacterium glutamicum strain with high efficiency of β-alanine production was constructed in this study. To do this, the biosynthetic pathway of β-alanine in an L-lysine-producing strain XQ-5 was modified by enhancing carbon flux in biosynthetic pathway and limiting carbon flux in competitive pathway. This study showed that replacement of L-aspartate kinase (AK) with wild-type AK and disruption of lactate dehydrogenase and alanine/valine aminotransferases increase β-alanine production because of decreasing the by-products accumulation. Moreover, L-aspartate-α-decarboxylase (ADC) from Bacillus subtilis was designed as the best enzyme for increasing β-alanine production, and its variant ( Bs ADC E56S/I88M ) showed the highest activity for catalyzing L-aspartate to generate β-alanine. To further increase β-alanine production, expression level of Bs ADC E56S/I88M was controlled by optimizing promoter and RBS, indicating that P gro plus ThirRBS is the best combination for Bs ADC E56S/I88M expression and β-alanine production. The resultant strain XQ-5.5 produced 30.7 ± 2.3 g/L of β-alanine with a low accumulation of lactate (from 5.2 ± 0.14 to 0.2 ± 0.09 g/L) and L-alanine (from 7.6 ± 0.22 to 3.8 ± 0. 32 g/L) in shake-flask fermentation and produced 56.5 ± 3.2 g/L of β-alanine with a productivity of 0.79 g/(L·h) and the glucose conversion efficiency (α) of 39.5% in feed-batch fermentation. This is the first report of genetically modifying the biosynthetic pathway of β-alanine that improves the efficiency of β-alanine production in an L-lysine-producing strain, and these results give us a new insight for constructing the other valuable biochemical. Key points • Optimization and overexpression of the key enzyme BsADC increased the accumulation of β-alanine. • The AK was replaced with wild-type AK to increase the conversion of aspartic acid to β-alanine. • A 56.5-g/L β-alanine production in fed-batch fermentation was achieved. Graphical abstract