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Won-Kyung Hong Sun-Yeon Heo Baek-Rock Oh Chul Ho Kim Jung-Hoon Sohn Ji-Won Yang Akihiko Kondo Jeong-Woo Seo 《Bioprocess and biosystems engineering》2013,36(9):1191-1197
In the present study, we established a genetic system for manipulating the oleaginous heterotrophic microalgae Aurantiochytrium sp. KRS101, using cycloheximide resistance as the selectable marker. The gene encoding ribosomal protein L44 (RPL44) of Aurantiochytrium sp. KRS101 was first identified and characterized. Proline 56 was replaced with glutamine, affording cycloheximide resistance to strains encoding the mutant protein. This resistance served as a novel selection marker. The gene encoding the Δ12-fatty acid desaturase of Mortierella alpina, used as a reporter, was successfully introduced into chromosomal DNA of Aurantiochytrium sp. KRS101 via 18S rDNA-targeted homologous recombination. Enzymatic conversion of oleic acid (C18:1) to linoleic acid (C18:2) was detected in transformants but not in the wild-type strain. 相似文献
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Won-Kyung Hong Chul-Ho Kim Sun-Yeon Heo Lian Hua Luo Baek-Rock Oh Dina Rairakhwada Jeong-Woo Seo 《Bioprocess and biosystems engineering》2011,34(2):231-236
Currently, 1,3-propanediol (1,3-PD) is an important chemical widely used in polymer production, but its availability is being
restricted owing to its expensive chemical synthesis. A methylotrophic yeast Hansenula polymorpha was engineered by expression of dhaB1, dhaB2, dhaB3, dhaB
RA1
and dhaB
RA2
encoding glycerol dehydratase complex and dhaT encoding 1,3-PD oxidoreductase from Klebsiella pneumoniae under direction of promoter of glyceraldehyde-3 phosphate dehydrogenase (GAPDH). The engineered recombinant yeast strain
can produce 1,3-PD from glucose (2.4 g L−1) as well as glycerol (0.8 g L−1), which might lead to a safe and cost-effective method for industrial production of 1,3-PD from various biomass resources. 相似文献
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Jang Min Park Baek-Rock Oh In Yeong Kang Sun-Yeon Heo Jeong-Woo Seo Seung-Moon Park Won-Kyung Hong Chul Ho Kim 《Journal of industrial microbiology & biotechnology》2017,44(7):1107-1113
A Bacillus sp. strain named BRC1 is capable of producing 2,3-butanediol (2,3-BD) using hydrolysates of the Jerusalem artichoke tuber (JAT), a rich source of the fructose polymer inulin. To enhance 2,3-BD production, we undertook an extensive analysis of the Bacillus sp. BRC1 genome, identifying a putative gene (sacC) encoding a fructan hydrolysis enzyme and characterizing the activity of the resulting recombinant protein expressed in and purified from Escherichia coli. Introduction of the sacC gene into Bacillus sp. BRC1 using an expression vector increased enzymatic activity more than twofold. Consistent with this increased enzyme expression, 2,3-BD production from JAT was also increased from 3.98 to 8.10 g L?1. Fed-batch fermentation of the recombinant strain produced a maximal level of 2,3-BD production of 28.6 g L?1, showing a high theoretical yield of 92.3%. 相似文献
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Lian Hua Luo Jeong-Woo Seo Sun-Yeon Heo Baek-Rock Oh Dae-Hyuk Kim Chul Ho Kim 《Bioprocess and biosystems engineering》2013,36(9):1319-1326
Klebsiella pneumoniae produces 3-hydroxypropionic acid (3-HP) from glycerol with oxidation of 3-hydroxypropionaldehyde (3-HPA) to 3-HP in a reaction catalyzed by aldehyde dehydrogenase (ALDH). In the present study, two putative ALDHs of K. pneumoniae, YneI and YdcW were identified and characterized. Recombinant YneI was specifically active on 3-HPA and preferred NAD+ as a cofactor, whereas YdcW exhibited broad substrate specificity and preferred NADP+ as a cofactor. Overexpression of ALDHs in the glycerol oxidative pathway-deficient mutant K. pneumoniae AK resulted in a significant increase in 3-HP production upon shake-flask culture. The final titers of 3-HP were 2.4 and 1.8 g L?1 by recombinants overexpressing YneI and YdcW, respectively. Deletion of the ALDH gene from K. pneumoniae did not affect the extent of 3-HP synthesis, implying non-specific activity of ALDHs on 3-HPA. The ALDHs might play major roles in detoxifying the aldehyde generated in glycerol metabolism. 相似文献
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Jang Min Park Won-Kyung Hong Sung-Mok Lee Sun-Yeon Heo You Ree Jung In Yeong Kang Baek-Rock Oh Jeong-Woo Seo Chul Ho Kim 《Journal of industrial microbiology & biotechnology》2014,41(9):1425-1433
Klebsiella pneumoniae synthesize large amounts of l-2,3-butanediol (l-2,3-BD), but the underlying mechanism has been unknown. In this study, we provide the first identification and characterization of an l-2,3-BD dehydrogenase from K. pneumoniae, demonstrating its reductive activities toward diacetyl and acetoin, and oxidative activity toward l-2,3-BD. Optimum pH, temperature, and kinetics determined for reductive and oxidative reactions support the preferential production of 2,3-BD during cell growth. Synthesis of l-2,3-BD was remarkably enhanced by increasing gene dosage, reaching levels that, to the best of our knowledge, are the highest achieved to date. 相似文献
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Luo LH Seo JW Baek JO Oh BR Heo SY Hong WK Kim DH Kim CH 《Applied microbiology and biotechnology》2011,89(3):697-703
Although the de novo biosynthetic mechanism of 3-hydroxypropionic acid (3-HP) in glycerol-fermenting microorganisms is still
unclear, the propanediol utilization protein (PduP) of Lactobacillus species has been suggested to be a key enzyme in this regard. To verify this hypothesis, a pduP gene from Lactobacillus reuteri was cloned and expressed, and the encoded protein was characterized. Recombinant L. reuteri PduP exhibited broad substrate specificity including 3-hydroxypropionaldehyde and utilized both NAD+ and NADP+ as a cofactor. Among various aldehyde substrates tested, the specific activity was highest for propionaldehyde, at pH 7.8
and 37 °C. The K
m and V
max values for propionaldehyde in the presence of NAD+ were 1.18 mM and 0.35 U mg−1, respectively. When L. reuteri pduP was overexpressed in Klebsiella pneumoniae, 3-HP production remarkably increased as compared to the wild-type strain (from 0.18 g L−1 to 0.72 g L−1) under shake-flask culture conditions, and the highest titer (1.38 g L−1 3-HP) was produced by the recombinant strain under batch fermentation conditions in a bioreactor. This is the first report
stating the enzymatic properties of PduP protein and the probable role in biosynthesis of 3-HP in glycerol fermentation. 相似文献
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Baek-Rock Oh Jeong-Woo Seo Min Ho Choi Chul Ho Kim 《Biotechnology and Bioprocess Engineering》2008,13(6):666-670
To produce 1,3-propanediol (1,3-PD) from crude glycerol, cultivation conditions were optimized by response surface methodology
(RSM) based on a 25 factorial central composite design (CCD). RSM was adopted to derive a statistical model for the individual and interactive
effects of crude glycerol, (NH4)2SO4, pH, cultivation time and temperature on the production of 1,3-PD. Optimal conditions for maximum 1,3-PD production were
as follows: crude glycerol, 35 g/L; (NH4)2SO4, 8 g/L; pH, 7.37; cultivation time, 10.8 h; temperature, 36.88°C. Under these optimal conditions, the design expert presented
the maximal numerical solution with a predicted 1,3-PD production level of up to 13.74 g/L. The experimental production of
1,3-PD yielded 13.8 g/L, which was in close agreement with the model prediction. 相似文献