Expression, Enzymatic Characterization, and High-Level Production of Glucose Isomerase from Actinoplanes missouriensis CICIM B0118(A) in Escherichia coli

2011 ◽  
Vol 66 (11-12) ◽  
pp. 605-613 ◽  
Author(s):  
He Wang ◽  
Ruijin Yang ◽  
Xiao Hua ◽  
Zhong Zhang ◽  
Wei Zhao ◽  
...  
Keyword(s):  
Ph Value ◽  
Glucose Isomerase ◽  
Medium Composition ◽  
Gene Encoding ◽  
E Coli ◽  
Optimum Ph ◽  
One Step ◽  
Ph Range ◽  
High Level ◽  
Level Production

High-level production of recombinant glucose isomerase (rGI) is desirable for lactulose synthesis. In this study, the xylA gene encoding glucose isomerase from Actinoplanes missouriensis CICIM B0118(A) was cloned and expressed in E. coli BL21(DE3), and high-level production was performed by optimization of the medium composition. rGI was purified from a recombinant E. coli BL21(DE3) and characterized. The optimum pH value of the purified enzyme was 8.0 and it was relatively stable within the pH range of 7.0 - 9.0. Its optimum temperature was around 85 °C, and it exhibited good thermostability when the temperature was lower than 90 °C. The maximum enzyme activity required the presence of both Co2+ and Mg2+, at the concentrations of 200 μM and 8 mM, respectively. With high-level expression and the simple one-step chromatographic purification of the His-tagged recombinant enzyme, this GI could be used in industrial production of lactulose as a potential economic tool

High-level production and one-step purification of biologically active human growth hormone in Escherichia coli

Gene ◽  
1995 ◽  
Vol 165 (2) ◽  
pp. 303-306 ◽  
Author(s):  
Reema Mukhija ◽  
Prithy Rupa ◽  
Devika Pillai ◽  
Lalit C. Garg
Keyword(s):  
Escherichia Coli ◽  
Growth Hormone ◽  
Human Growth Hormone ◽  
Human Growth ◽  
Biologically Active ◽  
One Step ◽  
High Level ◽  
Level Production

scholarly journals High-level production of membrane proteins in E. coli BL21(DE3) by omitting the inducer IPTG

2015 ◽  
Vol 14 (1) ◽  
Author(s):  
Zhe Zhang ◽  
Grietje Kuipers ◽  
Łukasz Niemiec ◽  
Thomas Baumgarten ◽  
Dirk Jan Slotboom ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
E Coli ◽  
High Level ◽  
Level Production

High-level production of soluble adenine nucleotide translocator from Schistosoma japonicum in E. coli cell-free system

2012 ◽  
Vol 47 (3) ◽  
pp. 395-400 ◽  
Author(s):  
Zhong Yang ◽  
Rongrong Ma ◽  
Lei Huang ◽  
Xiangcheng Zhu ◽  
Jiayuan Sheng ◽  
...  
Keyword(s):  
Schistosoma Japonicum ◽  
Adenine Nucleotide ◽  
Coli Cell ◽  
Adenine Nucleotide Translocator ◽  
Free System ◽  
Cell Free System ◽  
E Coli ◽  
High Level ◽  
Level Production

scholarly journals Antibiotic resistance and virulence of Escherichia coli strains isolated from animal rendering plant

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Gabriela Gregova ◽  
Vladimir Kmet
Keyword(s):  
Escherichia Coli ◽  
Antibacterial Agents ◽  
Virulence Genes ◽  
Gene Encoding ◽  
Antibiotic Resistant ◽  
Beta Lactamases ◽  
E Coli ◽  
Animal Wastes ◽  
Extended Spectrum Beta Lactamases ◽  
High Level

Abstract Processing of animal carcasses and other animal wastes in rendering plants is a significant source of antibiotic resistant microorganisms. The main goal of this study was to investigate the resistance to 18 antibacterial agents including β-lactams, fluoroquinolones, colistin and virulence factors (iss, tsh, cvaC, iutA, papC, kps and ibeA genes) in 88 Escherichia coli strains isolated from a rendering plant over 1 year period. ESBL (Extended-spectrum beta-lactamases) and plasmid-mediated Amp were screened by interpretative reading of MIC. ESBL phenotype was detected in 20.4% of samples and high level of resistance to fluoroquinolone was found in 27.2% of strains. Cephalosporinase CTX-M1, cephamycinase CMY-2, integrase 1 and transposon 3 genes were detected by PCR. Furthermore, there were found three CMY-2 producing E. coli with O25b-ST131, resistant to the high level of enrofloxacin and containing the gene encoding the ferric aerobactin receptor (iutA). One enrofloxacin resistant E. coli strain possessed iss, ibeA, kps and papC virulence genes also with CMY-2, integrase1 and Tn3. ST131 E. coli with CMY-2 has a zoonotic potential and presents a serious health risk to humans.

A New Operation of Carbohydrate-Containing Wastewater Treatment in an Anaerobic Fluidized Bed System

1992 ◽  
Vol 26 (9-11) ◽  
pp. 2453-2456 ◽  
Author(s):  
A. Matsumoto ◽  
M. Sakamoto ◽  
T. Noike
Keyword(s):  
Anaerobic Digestion ◽  
Fluidized Bed ◽  
Ph Value ◽  
Type Systems ◽  
Growth Type ◽  
Attached Growth ◽  
Optimum Ph ◽  
Effects Of Ph ◽  
Growth System ◽  
Ph Range

The effects of pH on the characteristics of conversion of starch to methane in anaerobic digestion were investigated by using a laboratory-scale anaerobic fluidized bed reactor over the pH range of 5.8-7.0. Furthermore, a new operation method was proposed to treat wastewater containing starch effectively. At pH 6.2, starch was decomposed well and methane was also formed well. The phenomenon showed methanogenesis in an attached growth system developed more tolerance for lower pH than in suspended growth type systems. After all, pH 6.2 was the optimum pH value for conversion of starch tomethane in our experiment. From these results, it is proposed that the more effective anaerobic digestion is made possible by the operation at low pH (for example 6.2) which accelerates acidogenesis and doesn't inhibit methanogenesis in attached growth systems.

Expression of β-agarase I DagA in prokaryotic cell and its activity identification

2009 ◽  
Vol 6 (2) ◽  
pp. 153-159
Author(s):  
Zhou Yan-Sheng ◽  
Wang Bao-Li ◽  
Qu Dong
Keyword(s):  
Molecular Chaperones ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Bacterial Protein ◽  
Gene Encoding ◽  
Sds Page ◽  
Optimum Ph ◽  
Pseudoalteromonas Atlantica ◽  
Polymerase Chain ◽  
Ph Range

AbstractThe DagA gene and DagA(▽), which is a DagA gene encoding sequence without signal peptide, were cloned from genome DNA ofPseudoalteromonas atlantica19262 by polymerase chain reaction (PCR). After ligation with pET21 vector, DagA and DagA(▽) were respectively expressed inEscherichia coliER2566 using molecular chaperones DsbC and FkpA. A strain of ER2566-pET21a-DagA(▽)-DsbC was screened as a highly effective expressing system in the form of an inclusion body that had the target protein with up to 60% total bacterial protein. DagA protein was renatured and purified by dissolving it in 8 mol/l of urea, using Ni-NTA resin affinity chromatography and refolding using the urea gradient method. DagA with a molecular weight of ~30.8 kDa was identified by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and had the ability to digest agarose. In a pH range of 4.8–6.8, DagA maintained a bioactivity greater than 60%, with 5.8 being the optimum pH, and it exhibited activity at temperatures from 37°C to 60°C, with 55°C being the optimum temperature.

scholarly journals High-level Production of Isoleucine and Fusel alcohol by expression of the Feedback Inhibition-insensitive Threonine deaminase in Saccharomyces cerevisiae

2022 ◽  
Author(s):  
Shota Isogai ◽  
Akira Nishimura ◽  
Atsushi Kotaka ◽  
Naoyuki Murakami ◽  
Natsuki Hotta ◽  
...  
Keyword(s):  
Parent Strain ◽  
Feedback Inhibition ◽  
Intracellular Accumulation ◽  
Threonine Deaminase ◽  
Gene Encoding ◽  
Acetate Ester ◽  
Fusel Alcohols ◽  
Fusel Alcohol ◽  
High Level ◽  
Level Production

A variety of the yeast Saccharomyces cerevisiae with intracellular accumulation of isoleucine (Ile) would be a promising strain for developing a distinct kind of sake, a traditional Japanese alcoholic beverage, because Ile-derived volatile compounds have a great impact on the flavor and taste of fermented foods. In this study, we isolated an Ile-accumulating mutant (strain K9-I48) derived from a diploid sake yeast of S. cerevisiae by conventional mutagenesis. Strain K9-I48 carries a novel mutation in the ILV1 gene encoding the His480Tyr variant of threonine deaminase (TD). Interestingly, the TD activity of the His480Tyr variant was markedly insensitive to feedback inhibition by Ile, but was not upregulated by valine, leading to intracellular accumulation of Ile and extracellular overproduction of 2-methyl-1-butanol, a fusel alcohol derived from Ile, in yeast cells. The present study demonstrated for the first time that the conserved histidine residue located in a linker region between two regulatory domains is involved in allosteric regulation of TD. Moreover, sake brewed with strain K9-I48 contained 2-3 times more 2-methyl-1-butanol and 2-methylbutyl acetate than sake brewed with the parent strain. These findings are valuable for the engineering of TD to increase the productivity of Ile and its derived fusel alcohols. IMPORTANCE Fruit-like flavors of isoleucine-derived volatile compounds, 2-methyl-1-butanol (2MB) and its acetate ester, contribute to a variety of the flavors and tastes of alcoholic beverages. Besides its value as aroma components in foods and cosmetics, 2MB has attracted significant attention as second-generation biofuels. Threonine deaminase (TD) catalyzes the first step in isoleucine biosynthesis and its activity is subject to feedback inhibition by isoleucine. Here, we isolated an isoleucine-accumulating sake yeast mutant and identified a mutant gene encoding a novel variant of TD. The variant TD exhibited much less sensitivity to isoleucine, leading to higher production of 2MB as well as isoleucine than the wild-type TD. Furthermore, sake brewed with a mutant yeast expressing the variant TD contained more 2MB and its acetate ester than that brewed with the parent strain. These findings will contribute to the development of superior industrial yeast strains for high-level production of isoleucine and its related fusel alcohols.

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