Ding R.,Jiangnan University |
Ding R.,the Key Laboratory of Industrial Biotechnology |
Li Z.,Jiangnan University |
Li Z.,the Key Laboratory of Industrial Biotechnology |
And 8 more authors.
Process Biochemistry | Year: 2010
The purpose of this study was to investigate the effect of medium additives on the secretion of recombinant α-cyclodextrin glucosyltransferase (α-CGTase) into the culture media of Escherichia coli. It is found that supplementation of the E. coli culture with SDS, glycine, Ca2+ or Na+, individually, facilitated the secretion of α-CGTase. Orthogonal experiment showed that the optimal condition to achieve maximal secretion of α-CGTase was the supplementation with 0.03% SDS, 400 mM Na+, 0.3% glycine and 10 mM Ca2+ together. Under this condition, extracellular enzyme activity reached 12.89 U/ml, which is 15 times higher than that of the culture without any additives. Further analysis showed that the permeability, fluidity and phosphatidylglycerol content of the E. coli cell membrane under the optimized condition were significantly increased in comparison to those under the control condition. These might be the potential mechanisms for the increased secretion of α-CGTase from the periplasmic compartment into the culture medium. © 2010 Elsevier Ltd. All rights reserved.
Guo Z.-P.,The Key Laboratory of Industrial Biotechnology |
Zhang L.,The Key Laboratory of Industrial Biotechnology |
Ding Z.-Y.,The Key Laboratory of Industrial Biotechnology |
Shi G.-Y.,The Key Laboratory of Industrial Biotechnology
Metabolic Engineering | Year: 2011
To synthesize glycerol, a major by-product during anaerobic production of ethanol, the yeast Saccharomyces cerevisiae would consume up to 4% of the sugar feedstock in typical industrial ethanol processes. The present study was dedicated to decreasing the glycerol production mostly in industrial ethanol producing yeast without affecting its desirable fermentation properties including high osmotic and ethanol tolerance, natural robustness in industrial processes. In the present study, the GPD1 gene, encoding NAD+-dependent glycerol-3-phosphate dehydrogenase in an industrial ethanol producing strain of S. cerevisiae, was deleted. Simultaneously, a non-phosphorylating NADP+-dependent glyceraldehyde-3-phosphate dehydrogenase (GAPN) from Bacillus cereus was expressed in the mutant deletion of GPD1. Although the resultant strain AG1A (gpd1Δ PPGK-gapN) exhibited a 48.7±0.3% (relative to the amount of substrate consumed) lower glycerol yield and a 7.6±0.1% (relative to the amount of substrate consumed) higher ethanol yield compared to the wild-type strain, it was sensitive to osmotic stress and failed to ferment on 25% glucose. However, when trehalose synthesis genes TPS1 and TPS2 were over-expressed in the above recombinant strain AG1A, its high osmotic stress tolerance was not only restored but also improved. In addition, this new recombinant yeast strain displayed further reduced glycerol yield, indistinguishable maximum specific growth rate (γmax) and fermentation ability compared to the wild type in anaerobic batch fermentations. This study provides a promising strategy to improve ethanol yields by minimization of glycerol production. © 2010 Elsevier Inc.