Abstract
Klebsiella pneumoniae KCTC2242 has high potential in the production of a high-value chemical, 2,3-butanediol (2,3-BDO). However, accumulation of metabolites such as lactate during cell growth prevent large-scale production of 2,3-BDO. Consequently, we engineered K. pneumoniae to redistribute its carbon flux toward 2,3-BDO production. The ldhA gene deletion and gene overexpression (budA and budB) were conducted to block a pathway that competitively consumes reduced nicotinamide adenine dinucleotide and to redirect carbon flux toward 2,3-BDO biosynthesis, respectively. These steps allowed efficient glucose conversion to 2,3-BDO under slightly acidic conditions (pH 5.5). The engineered strain SGSB105 showed a 40% increase in 2,3-BDO production from glucose compared with that of the host strain, SGSB100. Genes closely related to 2,3-BDO biosynthesis were observed at the gene transcription level by cultivating the SGSB100, SGSB103, SGSB104, and SGSB105 strains under identical growth conditions. Transcription levels for budA, budB, and budC increased approximately 10% during the log phase of cell growth relative to that of SGSB100. Transcription levels of 2,3-BDO genes in SGSB105 remained high during the log and stationary phases. Thus, the carbon flux was redirected toward 2,3-BDO production. Data on batch culture and gene transcription provide insight into improving the metabolic network for 2,3-BDO biosynthesis for industrial applications.
Highlights
Methods for engineering cell metabolism have enabled the production of high-value chemical compounds in large quantities
Among the five genes related to lactate biosynthesis, lactate dehydrogenase gene (ldhA) was identified by Macrogen Inc. (Seoul, South Korea) as a key gene for lactate production in K. pneumoniae KCTC2242
We constructed a pUC18K vector harboring the budA and budB genes and transported it into the SGSB100 strain to produce the SGSB104 strain. This approach was based on the effect of overexpression of budA, budB, and budC genes on 2,3-BDO biosynthesis reported in a study by Kim et al (2012)
Summary
Methods for engineering cell metabolism have enabled the production of high-value chemical compounds in large quantities. A single cell can be engineered to convert various carbon sources into valuable biochemical products such as lactate, butadiene, succinate, and 2,3-butanediol (2,3-BDO) [1,2]. Among these products, 2,3-BDO is essential for the synthesis of various biosynthetic products of economic value. As shown in previous studies, the conversion ratio of glucose into 2,3-BDO varies with pH and temperature [5,6,7]. Previous studies have confirmed that the enzyme activity of genes related to 2,3-BDO production is elevated at 37uC [10]. Even under optimized conditions for culture growth, the 2,3-BDO conversion ratio still cannot reach the optimal theoretical yield
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