• Proceedings of the Korean Society for Bioinformatics Conference
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• 2005.09a
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• pp.221-227
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• 2005

THEMATICS is a simple computational method for predicting functional sites in proteins. The method computes the theoretical titration curves of the ionizable residues of a protein using its 3D structure, determines the residues with perturbed, non-Henderson-Hasselbalch titration behavior, and identifies clusters of these perturbed residues in physical proximity. We have shown previously that this method is highly successful in predicting catalytic sites in enzymes. In the present study, we apply the method to non-catalytic ligand-binding proteins. It is shown that THEMATICS can predict non-catalytic binding sites. The success rate is better than 80 % for a set of 30 non-catalytic, ligand-binding proteins. The application of the method to Glutamine-binding protein from E. coli is discussed in detail.

• Journal of Microbiology and Biotechnology
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• v.15 no.6
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• pp.1267-1272
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• 2005

The effects of coexpression of GroEL/ES and DnaK/DnaJ/GrpE chaperones on the productivity of the soluble form of human granulocyte colony stimulating factor (hG-CSF) in E. coli were examined. Recombinant hG-CSF protein was coexpressed with DnaK/DnaJ/GrpE or GroEL/ES chaperones under the control of the araB or Pzt-1 promoter, respectively. The optimal concentration of L-arabinose for the expression of DnaK/DnaJ/GrpE was found to be 1 mg/ml. When L-arabinose was added at $OD_{600}$=0.2 (early-exponential phase), soluble hG-CSF production was greatly increased. In addition, it was observed that the DnaK/DnaJ/GrpE and GroEL/ES chaperones had no synergistic effects on preventing aggregation of hG-CSF protein. Consequently, by coexpression of the DnaK/DnaJ/GrpE chaperone, the signal intensity of the hG-CSF protein band in the soluble fraction of cell lysate was increased from $3.5\%\;to\;13.9\%$, and Western blot analysis also revealed about a 4-5-fold increase of production of soluble hG-CSF over the non-induction case of DnaK/DnaJ/GrpE.

• Journal of Microbiology and Biotechnology
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• v.9 no.5
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• pp.582-588
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• 1999

A Brevibacterium lactofermentum gene coding for a glucose-specific permease of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) was cloned, by complementing an Escherichia coli mutation affecting a ptsG gene with the B. lactofermentum genomic library, and completely sequenced. The gene was identified as a ptsG, which enables an E. coli transformant to transport non-metabolizable glucose analogue 2-deoxyglucose (2DG). The ptsG gene of B. lactofermentum consists of an open reading frame of 2,025 nucleotides encoding a polypeptide of 674 amino acid residues and a TAA stop codon. The 3' flanking region contains two stem-loop structures which may be involved in transcriptional termination. The deduced amino acid sequence of the B. lactofermentum enzyme $II^{GIe}$ specific to glucose ($EII^{GIe}$) has a high homology with the Corynebacterium glutamicum enzyme $II^{Man}$ specific to glucose and mannose ($EII^{Man}$), and the Brevibacterium ammoniagenes enzyme $II^{GIc}$ specific to glucose ($EII^{GIc}$). The 171-amino-acid C-terminal sequence of the $EII^{Glc}$ is also similar to the Escherichia coli enzyme $IIA^{GIc}$ specific to glucose ($IIA^{GIc}$). It is interesting that the arrangement of the structural domains, IIBCA, of the B. lactofermentum $EII^{GIc}$ protein is identical to that of EIIs specific to sucrose or $\beta$-glucoside. Several in vivo complementation studies indicated that the B. lactofermentum $EII^{Glc}$ protein could replace both $EII^{ Glc}$ and $EIIA^{Glc}$ in an E. coli ptsG mutant or crr mutant, respectively.

• Journal of Microbiology and Biotechnology
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• v.29 no.5
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• pp.776-784
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• 2019

Polyhydroxybutyrate (PHB), the most well-known polyhydroxyalkanoate, is a bio-based, biodegradable polymer that has the potential to replace petroleum-based plastics. Lignocellulose hydrolysate, a non-edible resource, is a promising substrate for the sustainable, fermentative production of PHB. However, its application is limited by the generation of inhibitors during the pretreatment processes. In this study, we investigated the feasibility of PHB production in E. coli in the presence of inhibitors found in lignocellulose hydrolysates. Our results show that the introduction of PHB synthetic genes (bktB, phaB, and phaC from Ralstonia eutropha H16) improved cell growth in the presence of the inhibitors such as furfural, 4-hydroxybenzaldehyde, and vanillin, suggesting that PHB synthetic genes confer resistance to these inhibitors. In addition, increased PHB production was observed in the presence of furfural as opposed to the absence of furfural, suggesting that this compound could be used to stimulate PHB production. Our findings indicate that PHB production using lignocellulose hydrolysates in recombinant E. coli could be an innovative strategy for cost-effective PHB production, and PHB could be a good target product from lignocellulose hydrolysates, especially glucose.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.2
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• pp.237-246
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• 2014

Two experiments were conducted to evaluate the efficacy of feeding an Escherichia coli (E. coli) derived phytase to pigs fed P deficient, corn-soybean meal diets. In Exp. 1, one hundred and twenty crossbred piglets ($9.53{\pm}0.84$ kg) were allocated to one of five treatments which consisted of four low P diets (0.61% Ca, 0.46% total P and 0.24% non-phytate P) supplemented with 0, 500, 1,000, or 20,000 FTU/kg E. coli phytase as well as a positive control formulated to be adequate in all nutrients (0.77% Ca, 0.62% total P and 0.42% non-phytate P). The treatments were applied to six pens with four pigs per pen for 28 days. In Exp. 2, ten crossbred pigs ($19.66{\pm}1.16$ kg) fitted with ileal T-cannula were used in a nutrient balance study. The pigs were assigned to treatments similar to those used in Exp. 1 in a doubly replicated $5{\pm}4$ incomplete Latin square design (5 diets with 4 periods). Each period consisted of a 5-d adjustment period followed by a 3-d total collection of feces and urine and then a 2-d collection of ileal digesta. Supplementation with phytase linearly increased (p<0.05) weight gain, feed intake, feed efficiency, bone breaking strength and fat-free dry and ash bone weight. There were linear increases (p<0.01) in the apparent ileal digestibility (AID) of DM, GE, CP, Ca, total P, inositol hexaphosphate ($IP_6$) and some AA with increasing dose of E. coli phytase. Pigs fed 20,000 FTU/kg had a greater (p<0.05) AID of IP6 (80% vs 59% or 64%, respectively) than pigs fed diets with 500 or 1,000 FTU/kg phytase. There were linear increases (p<0.05) in the total tract digestibility of Ca, total P, Na, K, Mg, and Zn as well as in the retention of Mg and Zn with increased phytase dose. The retention and utilization of Cu, and the total tract digestibility of CP and Cu quadratic increased (p<0.05) with increased phytase dose. In conclusion, supplementation of 500 FTU of phytase/kg and above effectively hydrolyzed phytate in low-P corn-soybean diets for pigs. In addition, a super dose of phytase (20,000 FTU/kg) hydrolyzed most of the IP6 and consequently further improved mineral use, protein utilization and performance.

• The Journal of the Korean Society for Microbiology
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• v.22 no.3
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• pp.233-240
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• 1987

The amniotic fluid provides a medium in which the fetus can readily move, cushions him against possible injury and helps him maintain an even temperature. Besides above mentioned functions, investigators reported that human amniotic fluid contains host-resistance factors which prevent bacteria from producing infectious disease and this activity shows difference among human racial groups or bacterial genera, species and strains. 40 amniotic fluid specimens from Korean women in their second and third trimesters of pregnancy were examined for inhibiting the growth of Escherichia coli. And various factors which might affect bacterial growth inhibiting activity such as pH, initial inoculum size, concentration of amniotic fluid, and heat resistance, were also tested using a strongly inhibitory amniotic fluid specimen. Finally plate diffusion tests were carried out using other strongly inhibitory amniotic fluid. The following results were obtained: 1. Of the 40 fluid samples examined, 18 specimens(45%) had inhibitory activity and samples from women in their second trimester of pregnanancy showed non-inhibitory activity(2 specimens). 2. The pH of the fluids varied between 7.43 and 8.33. There was no correlation between pH and inhibitory activity. 3. No. 19 amniotic fluid showed bacteriostatic activity after 24 hours incubation when an inoculum of $10^2$ organisms per milliliter was used, but non-inhibitory with an inoculum of $10^3$ and $10^4$ bacteria per milliliter. 4. The content of amniotic fluid in culture media influenced E. coli growth. At 90 percent, E. coli was inhibited growth but at 10 percent and 50 percent. 5. Inhibitory activity of No. 19 amniotic fluid was retained after heating to $50^{\circ}C$ for 30 minutes or 100^{\circ}C$ for 30 minutes. 6. Plate diffusion tests with No. 27 amniotic fluid showed that 0.7ml amniotic fluid gave clear zone of growth inhibition around the central well but 0.2ml and 0.1ml amniotic fluids were not.

• The Korean Journal of Pharmacology
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• v.16 no.2 s.27
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• pp.77-83
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• 1980

The effectiveness of Cefoperazone in the treatment of urinary tract infection was evaluated in the Department of Urology, St. Mary's hospital from December 1. 1979 to April 30. 1980. In this studies, the cusative organisms were divided into 2 groups; 1) Single form. E. coli (8), Staphylococcus aureus(7), Proteus vulgaris (4), P. morganii(3), S. epidermis (1), Enterococcus(1), Klebsiella (2), N. gonococcus(1). 2) Mixed from: Proteus+E. coli(4), E. coli+other(1), Pseudomonas+Enterococcus(1), Klebsiella+other(1). Effectiveness on urological diseases. 1) Neurogenic bladder: Results were excellent in 3 cases, good in 4 cases and negative effect in 4 cases. 2) Non-gonococcal urethritis: In this group, the therapeutic results were favorable in 88.9% of all cases. (Excellent in 2, Fail in1) 3) Pyelonephritis: All(4 cases) were excellent. 4) Renal stone: Among the 4 cases of renal stone, only one case was responded to cefoperazone. 5) Two cases of urethral stricture, two cases of cystitis, one case of B.P.H. and one case of gonococcal urethritis were all excellent. No serious side effects were observed except slight dizziness in one case.

• Korean Journal of Microbiology
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• v.40 no.2
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• pp.65-72
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• 2004

Instability of some eukaryotic sequence propagated in prokaryotic hosts is a frequently observed phenomenon. It is well documented that long inverted repeats, AT-rich sequences with structures like Z-DNA are extremely unstable in E. coli. These sequences may either be under-represented or even lost when cloned in E. coli. When we analyzed the polymorphic pattern for several tandom repeat (TR) in human SCKI gene, we found some TR regions were frequently deleted from plasmids and had difficult problem for their sequencing. These regions may result in non-clonability of the DNA sequence. Here we have cloned two difficult TR regions under low temperature and made two library for DNA sequencing using a nebulizer or sonicator. This study will help to determine the unstable genomic elements in complex mammalian genome.

• Magazine of the Korean Society of Agricultural Engineers
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• v.45 no.7
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• pp.94-106
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• 2003

A pilot study was performed to examine the feasibility of UV disinfection system and the reactivation of indicator microorganisms (TC, FC, E. coli) after UV irradiation for agricultural reuse of reclaimed water. Photoreactivation and dark repair enable UV-inactivated microorganisms to recover and may reduce the efficacy of UV inactivation, which might be drawbacks of the UV disinfection method. The effluent of biofilter for 16-unit apartment house was used as input to the UV disinfection system, and average SS and BOD concentration were 3.8 and 5.7 mg/L, respectively, and the mean level of total coliform was in the range of $1.0\times10^4$ MPN/100mL. UV disinfection was found to be effective and it reduced mean concentration of indicator microorganisms (total coliform, fecal coliform, and E. coli) to less than 100 MPN/100mL within 60s exposure using 17, 25, and 40W lamps. Two UV doses of 6 and 16 mW$\cdot$s/$\textrm{km}^2$ were applied and microorganisms reactivation was monitored under the dark, photoreactivating light, and solar irradiation. Microorganisms reactivation was observed in the UV dose of 6 mW$\cdot$s/$\textrm{km}^2$, and numbers increased up to 5% at the photoreactivating light and 1% at the dark. However, microorganisms were inactivated rather than reactivated at the solar radiation and numbers decreased to non-detectible level about below 2 MPN/100mL in 4 hours. In the case of 16 mW$\cdot$s/$\textrm{km}^2$, microorganism reactivation was not observed indicating that UV dose might affect the reactivation process such as photoreactivation and dark repair. Therefore, concerns associated with microorganism reactivation could be controlled by sufficient UV dose application. Agricultural reuse of reclaimed water might be even less concerned due to exposure to the solar irradiation that could further inactivate microorganisms. The pilot study result is encouraging, however, sanitary concern in water reuse is so critical that more comprehensive investigation is recommended.

• Microbiology and Biotechnology Letters
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• v.29 no.1
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• pp.1-11
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• 2001

Lactic acid bacteria (LAB) are widely used for various food fermentation. With the recent advances in modern biotechnology, a variety of bio-products with the high economic values have been produced using microorganisms. For molecular cloning and expression studies on the gene of interest, E. coli has been widely used mainly because vector systems are fully developed. Most plasmid vectors currently used for E, coli carry antibiotic-resistant markers. As it is generally believed that the antibiotic resistance markers are potentially transferred to other bacteria, application of the plasmid vectors carrying antibiotic resistance genes as selection markers should be avoided, especially for human consump-tion. By contrast, as LAB have some desirable traits such that the they are GRAS(generally recognized as safe), able to secrete gene products out of cell, and their low protease activities, they are regarded as an ideal organism for the genetic manipulation, including cloning and expression of homologous and heterologous genes. However, the vec-tor systems established for LAB are stil insufficient to over-produce gene products, stably, limiting the use of these organisms for industrial applications. For a past decade, the two popular plasmid vectors, pAM$\beta$1 of Streptococcus faecalis and pGK12 theB. subtilis-E. coli shuttle vector derived from pWV01 of Lactococcus lactis ssp. cremoris wg 2, were most widely used to construct efficient chimeric vectors to be stably maintained in many industrial strains of LAB. Currently, non-antibiotic markers such as nisin resistance($Nis^{r}$ ) are explored for selecting recombi-nant clone. In addition, a gene encoding S-layer protein, slp/A, on bacterial cell wall was successfully recombined with the proper LAB vectors LAB vectors for excretion of the heterologous gene product from LAB Many food-grade host vec-tor systems were successfully developed, which allowed stable integration of multiple plasmid copies in the vec-mosome of LAB. More recently, an integration vector system based on the site-specific integration apparatus of temperate lactococcal bacteriophage, containing the integrase gene(int) and phage attachment site(attP), was pub-lished. In conclusion, when various vector system, which are maintain stably and expressed strongly in LAB, are developed, lost of such food products as enzymes, pharmaceuticals, bioactive food ingredients for human consump-tion would be produced at a full scale in LAB.