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Phosphate-Responsive Promoter of a Pichia pastoris Sodium Phosphate Symporter
Ahn, Jungoh,Hong, Jiyeon,Park, Myongsoo,Lee, Hyeokweon,Lee, Eungyo,Kim, Chunsuk,Lee, Joohwan,Choi, Eui-sung,Jung, Joon-ki,Lee, Hongweon American Society for Microbiology 2009 Applied and environmental microbiology Vol.75 No.11
<B>ABSTRACT</B><P>To develop a functional phosphate-regulated promoter in <I>Pichia pastoris</I>, a phosphate-responsive gene, <I>PHO89</I>, which encodes a putative sodium (Na<SUP>+</SUP>)-coupled phosphate symporter, was isolated. Sequencing analyses revealed a 1,731-bp open reading frame encoding a 576-amino-acid polypeptide with 12 putative transmembrane domains. The properties of the <I>PHO89</I> promoter (<I>PPHO89</I>) were investigated using a bacterial lipase gene as a reporter in 5-liter jar fermentation experiments. <I>PPHO89</I> was tightly regulated by phosphate and was highly activated when the cells were grown in a phosphate-limited external environment. Compared to translation elongation factor 1α and the glyceraldehyde-3-phosphate dehydrogenase promoter, <I>PPHO89</I> exhibited strong transcriptional activity with higher specific productivity (amount of lipase produced/cell/h). Furthermore, a cost-effective and simple <I>PPHO89</I>-based fermentation process was developed for industrial application. These results demonstrate the potential for efficient use of <I>PPHO89</I> for controlled production of recombinant proteins in <I>P. pastoris</I>.</P>
Eun, Hyunmin,Kwon, Woo Young,Kalimuthu, Kalishwaralal,Kim, Yonghwan,Lee, Miran,Ahn, Jung-Oh,Lee, Hongweon,Lee, Sang Hyun,Kim, Hyung Joo,Park, Hyun Gyu,Park, Ki Soo The Royal Society of Chemistry 2019 Journal of Materials Chemistry B Vol.7 No.15
<P>A new method has been developed for the preparation of brightly fluorescent and stable DNA-silver nanoclusters (DNA-AgNCs). The approach takes advantage of specific interactions occurring between melamine and thymine residues in a DNA template. These interactions cause the formation of a melamine-DNA-AgNC complex (Mel-DNA-AgNCs), in which a change in the environment of the DNA template causes binding of additional Ag<SUP>+</SUP> and an enhancement in the fluorescence efficiency and stability. The effects of the nature of the template DNA, DNA : Ag<SUP>+</SUP> : NaBH4 ratio, pH and temperature were systematically assessed in order to maximize the melamine-promoted fluorescence enhancement. The results show that the Mel-DNA-AgNCs, generated under the optimal conditions, exhibit a <I>ca.</I> 3-fold larger fluorescence efficiency and long-term stability (70 d) in contrast to those of DNA-AgNCs in the absence of melamine. Importantly, the bright and stable Mel-DNA-AgNCs exhibit antimicrobial activities against Gram-positive and Gram-negative bacteria that are superior to those of DNA-AgNCs alone. To the best of our knowledge, this is the first report describing the synthesis of DNA-AgNCs that have improved fluorescence efficiencies and that function as effective antimicrobial agents.</P>
Seo, Hyung-Min,Jeon, Jong-Min,Lee, Ju Hee,Song, Hun-Suk,Joo, Han-Byul,Park, Sung-Hee,Choi, Kwon-Young,Kim, Yong Hyun,Park, Kyungmoon,Ahn, Jungoh,Lee, Hongweon,Yang, Yung-Hun Springer-Verlag 2016 Journal of industrial microbiology & biotechnology Vol.43 No.1
<P>Furfural is a toxic by-product formulated from pretreatment processes of lignocellulosic biomass. In order to utilize the lignocellulosic biomass on isobutanol production, inhibitory effect of the furfural on isobutanol production was investigated and combinatorial application of two oxidoreductases, FucO and YqhD, was suggested as an alternative strategy. Furfural decreased cell growth and isobutanol production when only YqhD or FucO was employed as an isobutyraldehyde oxidoreductase. However, combinatorial overexpression of FucO and YqhD could overcome the inhibitory effect of furfural giving higher isobutanol production by 110 % compared with overexpression of YqhD. The combinatorial oxidoreductases increased furfural detoxification rate 2.1-fold and also accelerated glucose consumption 1.4-fold. When it compares to another known system increasing furfural tolerance, membrane-bound transhydrogenase (pntAB), the combinatorial aldehyde oxidoreductases were better on cell growth and production. Thus, to control oxidoreductases is important to produce isobutanol using furfural-containing biomass and the combinatorial overexpression of FucO and YqhD can be an alternative strategy.</P>
Ahn, Jungoh,Chung, Bevan K S,Lee, Dong-Yup,Park, Myongsoo,Karimi, Iftekhar A,Jung, Joon-Ki,Lee, Hongweon Published by Elsevier/North Holland on behalf of t 2011 FEMS microbiology letters Vol.324 No.1
<P>We explored the physiological and metabolic effects of different carbon sources (glucose, fructose, and glucose/fructose mixture) in phosphoglucose isomerase (pgi) knockout Escherichia coli mutant producing shikimic acid (SA). It was observed that the pgi(-) mutant grown on glucose exhibited significantly lower cell growth compared with the pgi(+) strain and its mixed glucose/fructose fermentation grew well. Interestingly, when fructose was used as a carbon source, the pgi(-) mutant showed the enhanced SA production compared with the pgi(+) strain. In silico analysis of a genome-scale E. coli model was then conducted to characterize the cellular metabolism and quantify NAPDH regeneration, which allowed us to understand such experimentally observed attenuated cell growth and enhanced SA production in glucose- and fructose-consuming pgi(-) mutant, respectively with respect to cofactor regeneration.</P>
Kim, Young Su,Yoon, Nam-kyung,Karisa, Nadia,Seo, Sin-hye,Lee, Jeong-soo,Yoo, Sung-sik,Yoon, In-joong,Kim, Yeu-chun,Lee, Hongweon,Ahn, Jungoh Academic Press 2019 Microbial pathogenesis Vol.127 No.-
<P><B>Abstract</B></P> <P> <I>Streptococcus parauberis</I> is the major infectious agent of streptococcosis in the olive flounder (<I>Paralichthys olivaceus</I>), causing serious economic damage. In this study, we identified potential vaccine candidates against <I>S. parauberis</I> by reverse vaccinology. In total, the 2 out of 21 proteins were identified as vaccine candidates from two available <I>S. parauberis</I> genomes. The membrane-anchored protein SEC10/PgrA and the metal ABC transporter substrate-binding lipoprotein mtsA were potent antigenic proteins based on western blotting with mouse-derived antiserum against whole bacteria of <I>S. parauberis</I> serotypes I and II. In particular, metal ABC transporter substrate-binding lipoprotein (<I>mtsA</I>) showed similar protective immunity to that of whole-cell bacterins against <I>S. parauberis</I> in a zebrafish model. These results suggest that mtsA may be considered as a novel candidate in the development of vaccines against <I>S. parauberis</I>.</P> <P><B>Highlights</B></P> <P> <UL> <LI> We identified 41 vaccine candidates against <I>Streptococcus parauberis.</I> </LI> <LI> SEC10/PgrA and mtsA were potent antigenic proteins. </LI> <LI> <I>mtsA</I> showed high protective immunity in a zebrafish model. </LI> </UL> </P>
Hong, Minhee,Ahn, Jungoh,Yoo, Sungsik,Hong, Jiyeon,Lee, Eungyu,Yoon, Injoong,Jung, Joon-ki,Lee, Hongweon Elsevier 2011 Veterinary microbiology Vol.148 No.1
<P><B>Abstract</B></P><P><I>Haemophilus parasuis</I> causes contagious porcine Glässer's disease, which is occurring worldwide and leads to severe losses in the pig industry. To identify novel antigen candidates against this disease, 22 surface-exposed or secreted proteins were selected from the annotated <I>H. parasuis</I> genome by reverse vaccinology strategy. Expression of these proteins in <I>Escherichia coli</I> was attempted. Immunogenicity of the expressed candidates was assessed using Western blot analysis with mouse-derived antiserum prepared with whole bacteria of <I>H. parasuis</I> serovar 4 or 5. Three ABC-type transporters (OppA, YfeA and PlpA) and 1 curli protein assembly (CsgG) were identified as potent immunogenic proteins. The proteins show cross-reactions when tested with sera raised against serovars 4 and 5 of <I>H. parasuis</I>.</P>
High-level recombinant production of squalene using selected Saccharomyces cerevisiae strains
Han, Jong Yun,Seo, Sung Hwa,Song, Jae Myeong,Lee, Hongweon,Choi, Eui-Sung Springer-Verlag 2018 Journal of industrial microbiology & biotechnology Vol.45 No.4
<P>For recombinant production of squalene, which is a triterpenoid compound with increasing industrial applications, in microorganisms generally recognized as safe, we screened Saccharomyces cerevisiae strains to determine their suitability. A strong strain dependence was observed in squalene productivity among Saccharomyces cerevisiae strains upon overexpression of genes important for isoprenoid biosynthesis. In particular, a high level of squalene production (400 +/- 45 mg/L) was obtained in shake flasks with the Y2805 strain overexpressing genes encoding a bacterial farnesyl diphosphate synthase (ispA) and a truncated form of hydroxyl-3-methylglutaryl-CoA reductase (tHMG1). Partial inhibition of squalene epoxidase by terbinafine further increased squalene production by up to 1.9-fold (756 +/- 36 mg/L). Furthermore, squalene production of 2011 +/- 75 or 1026 +/- 37 mg/L was obtained from 5-L fed-batch fermentations in the presence or absence of terbinafine supplementation, respectively. These results suggest that the Y2805 strain has potential as a new alternative source of squalene production.</P>
Jeon, Wooyoung,Kim, Yeu-Chun,Hong, Minhee,Rejinold, Sanoj,Park, Kyoungmoon,Yoon, Injoong,Yoo, Sungsik,Lee, Hongweon,Ahn, Jungoh Hindawi 2018 BioMed research international Vol.2018 No.-
<P>The study describes the development of a vaccine using microcrystalline cellulose (Avicel PH-101) as a delivery carrier of recombinant protein-based antigen against erysipelas. Recombinant SpaA, surface protective protein, from a gram-positive pathogen<I> Erysipelothrix rhusiopathiae</I> was fused to a cellulose-binding domain (CBD) from<I> Trichoderma harzianum</I> endoglucanase II through a S3N10 peptide. The fusion protein (CBD-SpaA) was expressed in<I> Escherichia coli</I> and was subsequently bound to Avicel PH-101. The antigenicity of CBD-SpaA bound to the Avicel was evaluated by enzyme-linked immunosorbent (ELISA) and confocal laser scanning microscope (CLSM) assays. For the examination of its immunogenicity, groups of mice were immunized with different constructs (soluble CBD-SpaA, Avicel coated with CBD-SpaA, whole bacterin of<I> E. rhusiopathiae</I> (positive control), and PBS (negative control)). In two weeks after immunization, mice were challenged with 1x10<SUP>7</SUP> CFU of<I> E. rhusiopathiae</I> and Avicel coated with CBD-SpaA induced protective immunity in mice. In conclusion, this study demonstrates the feasibility of microcrystalline cellulose as the delivery system of recombinant protein subunit vaccine against<I> E. rhusiopathiae </I>infection in mice.</P>