• Asian-Australasian Journal of Animal Sciences
• /
• 제25권9호
• /
• pp.1255-1261
• /
• 2012

Thirty two Holstein female calves (initial body weight = $40{\pm}3.0$ kg) were used to investigate the effects of probiotic and prebiotic on average daily gain (ADG), fecal E. coli count, white blood cell count, plasma IgG1 level and cell-mediated immune response to injection of phytohemagglutinin in suckling female calves. Calves were assigned randomly to one of the four treatments, including whole milk without additives (control), whole milk containing probiotic, whole milk containing prebiotic and whole milk containing probiotic and prebiotic (synbiotic). Average daily gain was greater in calves fed probiotic, prebiotic and synbiotic at weeks 6, 7 and 8 (p<0.05). E. coli count was significantly lower in calves fed probiotic, prebiotic and synbiotic on d 56 (p<0.05). There was no significant difference between treatments in blood samples and cell-mediated response. This study showed that addition of probiotic, prebiotic and combination of these additives to milk enhanced ADG and reduced fecal E. coli count in preruminant calves.

• 한국미생물생명공학회:학술대회논문집
• /
• 한국미생물생명공학회 2000년도 Proceedings of 2000 KSAM International Symposium and Spring Meeting
• /
• pp.68-75
• /
• 2000

D-Tagatose is a potential bulking agent in food as a non-calorific sweetener. To produce D-tagatose from cheaper resources, plasmids harboring the L-arabinose isomerase gene (araA) from Escherichia coli was constructed because L-arabinose isomerase was previously suggested as an enzyme that mediates the bioconversion of galactose to tagatose as well as that of arabinose to ribulose. In the cultures of recombinant E.coli with pTC101, which harboring araA of E.coli, tagatose was produced from galactose in 9.9 % yield. The enzyme extract of E.coli containing pTC101 also converted galactose into tagatose in 96.4 % yield. For the economic production of D-tagatose, an L-arabinose isomerase of E.coli was immobilized using covalent binding on agarose. While the free L-arabinose isomerase produced tagatose with the rate of 0.48 mg/U$.$day, the immobilized one stably converted galactose into average 7.5 g/l$.$day of tagatose during 7 days with higher productivity of 0.87 mg/U$.$day. In the scaled up immobilized enzyme system, 99.9 g/l of tagatose was produced from galactose with 20 % equilibrium in 48 hrs. The process was stably repeated additional 2 times with tagatose production of 104.1 and 103.5 g/l.

• Journal of Microbiology and Biotechnology
• /
• 제13권2호
• /
• pp.287-291
• /
• 2003

Botulinum neurotoxin type A (BONT/A) is an extremely potent toxin, which is produced by Clostridium botulinum. The light chain of this protein (BONT/A LC), which is known as a zinc endopeptidase, cleaves SNAP-25 involved in the exocytosis process. In this work, the expression of recombinant BoNT/A LC in E. coli is described. The BONT/A LC gene of C. botulinum contains a high frequency of the arginine AGA and isoleucine ATA codons that are rarely used in genes of E. coli, hampering the translation of recombinant protein. The argD and ilex tRNA genes were cloned into pACYC184 vector, resulting in pAAD131X plasmid. The translational stress of the toxin gene related to codon bias was reversed by fupplernentation of the AGA arginyl tRNA of T4 phage and AUA isoleucyl tRNA of E. coli. This system may be applicable for the expression of a variety of AT-rich heterologous genes in E. coli.

• Molecules and Cells
• /
• 제28권4호
• /
• pp.397-401
• /
• 2009

Flavonoids are a group of polyphenolic compounds that have been recognized as important due to their physiological and pharmacological roles and their health benefits. Glycosylation of flavonoids has a wide range of effects on flavonoid solubility, stability, and bioavailability. We previously generated the E. coli BL21 (DE3) ${\Delta}pgi$ host by deleting the glucose-phosphate isomerase (Pgi) gene in E. coli BL21 (DE3). This host was further engineered for whole-cell biotransformation by integration of galU from E. coli K12, and expression of calS8 (UDP-glucose dehydrogenase) and calS9 (UDP-glucuronic acid decarboxylase) from Micromonospora echinospora spp. calichensis and arGt-4 (7-O-glycosyltransferase) from Arabidopsis thaliana to form E. coli (US89Gt-4), which is expected to produce glycosylated flavonoids. To test the designed system, the engineered host was fed with naringenin as a substrate, and naringenin 7-O-xyloside, a glycosylated naringenin product, was detected. Product was verified by HPLC-LC/MS and ESI-MS/MS analyses. The reconstructed host can be applied for the production of various classes of glycosylated flavonoids.

• Journal of Microbiology and Biotechnology
• /
• 제22권2호
• /
• pp.234-238
• /
• 2012

Recombinant Escherichia coli displaying organophosphorus hydrolase (OPH) was used to overcome the diffusion barrier limitation of organophosphorus pesticides. A new anchor system derived from the N-terminal domain of ice-nucleation protein from Pseudomonas syringae InaV (InaV-N) was used to display OPH onto the surface. The designed sequence was cloned in the vector pET-28a(+) and then was expressed in E. coli. Tracing of the expression location of the recombinant protein using SDS-PAGE showed the presentation of OPH by InaV-N on the outer membrane, and the ability of recombinant E. coli to utilize diazinon as the sole source of energy, without growth inhibition, indicated its significant activity. The location of OPH was detected by comparing the activity of the outer membrane fraction with the inner membrane and cytoplasm fractions. Studies revealed that recombinant E. coli can degrade 50% of 2 mM chlorpyrifos in 2 min. It can be concluded that InaV-N can be used efficiently to display foreign functional protein, and these results highlight the high potential of an engineered bacterium to be used in bioremediation of pesticide-contaminated sources in the environment.

• 한국동물위생학회지
• /
• 제44권4호
• /
• pp.247-256
• /
• 2021

The recombinant lysozyme-HJL34 proteins were expressed and purified using commercial Escherichia (E.) coli expression system. Stx2e⁺ F18⁺ E. coli, Actinobacillus pleuropneumoniae (APP), Streptococcus (S.) suis, and Clostridium (C.) perfringens strains were isolated from pigs. The minimum inhibitory concentrations (MICs) of the recombinant lysozyme-HJP34 proteins were examined by means of the microtiter plate method, according to the NCCLS recommendations. The possibility of its as the alternatives to antibiotics was tested in piglets. The MICs were determined as 75 ㎍/mL, 300 ㎍/mL, 75 ㎍/mL, 35.5 ㎍/m against Stx2e⁺ F18⁺ E. coli, APP, S. suis, C. perfringens, respectively. A total of 25 piglets were divied 5 groups. The piglets in group A~C were fed with commercial feed and those in groups D, E were fed with commercial feedstuff. All piglets in groups B~E were challenged with virulent Stx2e⁺ F18⁺ E. coli, APP, S. suis strains. Groups C and D were treated with antimicrobial from 24 h after challenge. All piglets in group B died within 3 days after challenge. Among 5 piglets in groups C and D piglets, 80% survived after challenge. Among group E piglets, 60% were alive until the end of this study. Therefore, this study indicates that recombinant lysozyme-HJP34 proteins is a suitable possibility as a feed additive for reduction of diseases by bacterial pathogens in piglet feed.

• KSBB Journal
• /
• 제19권4호
• /
• pp.327-330
• /
• 2004

Inducible system을 이용하여 (R)-hydroxybutyric acid (R3HB)를 생산하는 재조합 대장균을 개발하였다. Ralstonia eutropha에서 유래한 PHB depolymerase 유전자를 inducible promoter에 의하여 발현되게 만들고, PHB 생합성 유전자가 있는 vector에 cloning하였다. PHB 생합성 유전자와 depolymerase 유전자를 가지고 있는 재조합 대장균을 배양하여 먼저 PHB를 축적시킨 후에 depolymerase를 발현시켜 배지 내로 분비된 R3HB를 확인하였다. 그 결과 축적된 PHB의 대부분은 발현된 depolymerase에 의하여 분해되었으며, depolymerase의 발현 이후에도 재조합 대장균은 PHB를 축적하고 분해하여 R3HB의 농도는 배양시간에 따라 증가하였다. 이러한 결과는 inducible depolymerase를 갖는 재조합 대장균을 이용하여 높은 농도의 R3IB를 얻을 수 있다는 것을 보여준다.

• Journal of Microbiology and Biotechnology
• /
• 제8권3호
• /
• pp.214-221
• /
• 1998

A Brevibacterium ammoniagenes gene coding for glucose/mannose-specific enzyme II ($EII^{Glc}$) of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) was cloned by complementing an Escherichia coli mutation affecting a ptsG gene, and the complete DNA nucleotide sequence was determined. The cloned gene was identified to be a ptsG, which enables the E. coli transportment to use glucose more efficiently than mannose as the sole carbon source in an M9 minimal medium. The ptsG gene of B. ammoniagenes consists of an open reading frame of 1,983 nucleotides putatively encoding a polypeptide of 661 amino acid residues and a TAA stop codon. The deduced amino acid sequence of the B. ammoniagenes $EII^{Glc}$ shows, at $46\%$, the highest degree of sequence similarity with the Corynebacterium glutamicum EII specific for both glucose and mannose. In addition, the $EII^{Glc}$ shares approximately $30\%$ sequence similarities with sucrose-specific and ${\beta}$-glucoside-specific EIIs of the several bacteria belonging to the glucose-PTS class. The 161-amino-acid C-terminal sequence of $EII^{Glc}$ is also similar to that of E. coli enzyme $IIA^{Glc}$, specific for glucose ($EIIA^{Glc}$). The B. ammoniagenes $EII^{Glc}$ consists of three domains; a hydrophobic region (EIIC) and two hydrophilic regions (EIIA, EIIB). The arrangement of structural domains, IIBCA, of the $EII^{Glc}$ is identical to those of EIIs specific for sucrose or ${\beta}$-glucoside. While the domain IIA was removed from the B. ammoniagenes $EII^{Glc}$ the remaining domains IIBC were found to restore the glucose and mannose-utilizing capacity of E. coli mutant lacking $EII^{Glc}$ activity with $EIIA^{Glc}$ of the E. coli mutant. $EII^{Glc}$ contains a histidine residue and a cysteine residue which are putative phosphorylation sites for the protein.

• Journal of Electrical Engineering and Technology
• /
• 제6권2호
• /
• pp.275-279
• /
• 2011

Sterilization of Escherichia coli (E. coli) is examined using a unique pulsed ultra-violet (UV) elliptical reactor based on Nd:YAG laser resonator, UV radiation from a pulsed xenon flashlamp. The light from the discharge has a broadband emission spectrum extending from the UV to the infrared region with a rich UV contained. Sterilization method by using the UV light is fast, environment-friendly and it does not cause secondary pollution. A Nd:YAG laser resonator having elliptical shape has advantage of concentrating the radiation of the UV light at two foci as the quart sleeve filled with E. coli. The primary objective of this research is to determine the important parameters such as pulse per second (pps), the applied voltage for sterilizing E. coli by using an UV elliptical reactor. From the experiment result, the sterilization effect of UV elliptical reactor is better than that of UV cylindrical reactor, and it can be 99.9% of sterilization at 800V regardless of the pps within 10 minutes.

• Archives of Pharmacal Research
• /
• 제21권3호
• /
• pp.305-309
• /
• 1998

Human cytochrome P450 1A2 is one of the major cytochrome P450s in human liver. It is known to be capable of activating a number of carcinogens such as arylamines and heterocyclic amines. In order to develop the new bacterial mutagenicity test system with human P450, a full length of human P450 1A2 cDNA inserted into pCW bacterial expression vector was introduced to Escherichia coli WP2 uvrA strain which is a well-known E. coli strain for bacterial reverse mutagenicity assay. Expressed human P450 1A2 showed typical P450 hemoprotein spectra. Maximum expression was achieved at 48 hrs after incubating at $30^{\circ}C$ in terrific broth containing ampicillin, IPTG and other supplements. High level expression of P450 1A2 in E. coli WP2 uvrA membranes was determined in SDS-PAGE. The well-known mutagens 2-aminoanthracene and MElQ increased the revertant colonies of E. coli WP2 uvrA expressing human P450 1A2 without an exogenous rat hepatic post-mitochondrial supernatant (S9 fraction) in a dose-dependent manner. The results show that the functional expression of human P450 in bacterial mutagenicity tester strain will provide a useful tool for studying the mechanism of the mutagenesis and carcinogenesis of new drugs and environmental chemicals.