Dina Rairakhwada

Identification and utilization of a 1,3-propanediol oxidoreductase isoenzyme for production of 1,3-propanediol from glycerol in Klebsiella pneumoniae

In a previous study, we showed that 1,3-propanediol (1,3-PD) was still produced from glycerol by the Klebsiella pneumoniae mutant strain defective in 1,3-PD oxidoreductase (DhaT), although the production level was lower compared to the parent strain. As a potential candidate for another putative 1,3-PD oxidoreductase, we identified and characterized a homolog of Escherichia coli yqhD (88% homology in amino acid sequence), which encodes an alcohol dehydrogenase and is well known to replace the function of DhaT in E. coli. Introduction of multiple copies of the yqhD homolog restored 1,3-PD production in the mutant K. pneumoniae strain defective in DhaT. In addition, by-product formation was still eliminated in the recombinant strain due to the elimination of the glycerol oxidative pathway. An increase in NADP-dependent 1,3-PD oxidoreductase activity was observed in the recombinant strain harboring multiple copies of the yqhD homolog. The level of 1,3-PD production during batch fermentation in the recombinant strain was comparable to that of the parent strain; further engineering can generate an industrial strain producing 1,3-propanediol.

Growth of the oleaginous microalga Aurantiochytrium sp. KRS101 on cellulosic biomass and the production of lipids containing high levels of docosahexaenoic acid

We examined the growth of a novel oleaginous microalga, Aurantiochytrium sp. KRS101, using cellulosic materials as nutrients, and the resultant production of lipids containing high levels of docosahexaenoic acid (DHA). The microalgal strain could grow using either carboxymethylcellulose or cellobiose as a carbon source, and produced lipids containing high levels of DHA (49–58% of total fatty acids). In line with this growth behavior, carboxymethylcellulase and cellobiohydrolase activities were evident in both cell-free lysates and culture broths. Additionally, an industrial cellulosic biomass, palm oil empty fruit bunches (POEFB), a by-product of the palm oil industry, were utilized by the microalgal strain for cell growth and lipid production.

An efficient plasmid vector for constitutive high-level expression of foreign genes in Escherichia coli

The levansucrase gene (lsrA) from Rahnella aquatilis was strongly expressed in a constitutive manner in Escherichia coli when cloned into a pBluescript KS-based pRL1CP plasmid vector. The native promoter upstream of lsrA and the lacZ promoter cooperatively enhanced the expression of lsrA to a level that was comparable to that of the T7 promoter, which is used in commercial pET expression vector system. A putative rho-independent transcription termination signal downstream of lsrA was crucial for gene expression. This plasmid vector also proved to be applicable for efficient expression of other foreign genes in E. coli.