Alanine production in an H+-ATPase- and lactate dehydrogenase-defective mutant of Escherichia coli expressing alanine dehydrogenase.
Previously, we reported that pyruvate production was markedly improved in TBLA-1, an H(+)-ATPase-defective Escherichia coli mutant derived from W1485lip2, a pyruvate-producing E. coli K-12 strain. TBLA-1 produced more than 30 g/l pyruvate from 50 g/l glucose by jar fermentation, while W1485lip2 produced only 25 g/l pyruvate (Yokota et al. in Biosci Biotechnol Biochem 58:2164-2167, 1994b). In this study, we tested the ability of TBLA-1 to produce alanine by fermentation. The alanine dehydrogenase (ADH) gene from Bacillus stearothermophilus was introduced into TBLA-1, and direct fermentation of alanine from glucose was carried out. However, a considerable amount of lactate was also produced. To reduce lactate accumulation, we knocked out the lactate dehydrogenase gene (ldhA) in TBLA-1. This alanine dehydrogenase-expressing and lactate dehydrogenase-defective mutant of TBLA-1 produced 20 g/l alanine from 50 g/l glucose after 24 h of fermentation. The molar conversion ratio of glucose to alanine was 41%, which is the highest level of alanine production reported to date. This is the first report to show that an H(+)-ATPase-defective mutant of E. coli can be used for amino acid production. Our results further indicate that H(+)-ATPase-defective mutants may be used for fermentative production of various compounds, including alanine.
Wada
Masaru
M
Division of Applied Bioscience, Research Faculty of Agriculture, Hokkaido University, Kita-9, Nishi-9, Kita-ku, Sapporo, 060-8589, Japan. wada@chem.agr.hokudai.ac.jp
Narita
Kotomi
K
Yokota
Atsushi
A
D000409:Alanine / Q000096:biosynthesis*; D001709:Biotechnology / Q000379:methods*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005285:Fermentation; D007770:L-Lactate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D016254:Mutagenesis, Insertional; D006180:Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*
D004269:DNA, Bacterial; D007770:L-Lactate Dehydrogenase; D006180:Proton-Translocating ATPases; D000409:Alanine