Expression of the Klebsiella pneumoniae CG21 acetoin reductase gene in Clostridium acetobutylicum ATCC 824

• Published in: Journal of industrial microbiology & biotechnology 2001-10, Vol.27 (4), p.220-227
• Main Authors: WARDWELL, S. A, YANG, Y. T, CHANG, H. Y, SAN, K. Y, RUDOLPH, F. B, BENNETT, G. N
• Format: Article
• Language: English
• Subjects: 2,3-Butanediol; 2,3-Butylene glycol; Acetoin - metabolism; acetoin reductase; acetone reductase; Alcohol Oxidoreductases - genetics; Alcohol Oxidoreductases - metabolism; Amino Acid Sequence; Base Sequence; Biochemistry; Biological and medical sciences; Biotechnology; Butylene Glycols - metabolism; Cloning; Cloning, Molecular; Clostridium - enzymology; Clostridium - genetics; Clostridium - growth & development; Clostridium acetobutylicum; Culture Media; DNA methylation; E coli; Enzymes; Escherichia coli - enzymology; Escherichia coli - genetics; Escherichia coli - growth & development; Fermentation; Fundamental and applied biological sciences. Psychology; Gas chromatography; Genes; Genetic engineering; Genetic technics; Industrial production; Klebsiella pneumoniae; Klebsiella pneumoniae - enzymology; Klebsiella pneumoniae - genetics; Metabolism; Methods. Procedures. Technologies; Microbiology; Microorganisms; Modification of gene expression level; Molecular Sequence Data; Plasmids; Sequence Analysis, DNA; Solvents
• ISSN: 1367-5435; 1476-5535
• DOI: 10.1038/sj.jim.7000179

Acetoin reductase catalyzes the production of 2,3-butanediol from acetoin. The gene encoding the acetoin reductase of Klebsiella pneumoniae CG21 was cloned and expressed in Escherichia coli and Clostridium acetobutylicum ATCC 824. The nucleotide sequence of the gene encoding the enzyme was determined to be 768 bp long. Expression of the K. pneumoniae acetoin reductase gene in E. coli revealed that the enzyme has a molecular mass of about 31,000 Da based on sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis. The K. pneumoniae acetoin reductase gene was cloned into a clostridial/E. coli shuttle vector, and expression of the gene resulted in detectable levels of acetoin reductase activity in both E. coli and C. acetobutylicum. While acetoin, the natural substrate of acetoin reductase, is a typical product of fermentation by C. acetobutylicum, 2,3-butanediol is not. Analysis of culture supernatants by gas chromatography revealed that introduction of the K. pneumoniae acetoin reductase gene into C. acetobutylicum was not sufficient for 2,3-butanediol production even though the cultures were producing acetoin. 2,3-Butanediol was produced by cultures of C. acetobutylicum containing the gene only when commercial acetoin was added.