• Bulletin of the Korean Chemical Society
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• v.34 no.8
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• pp.2419-2424
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• 2013

Guanosine-5'-diphosphate 3'-diphosphate (ppGpp) serves as alarmone in bacterial stringent responses. In this study, an affinity column was constructed by immobilizing ppGpp to NHS-Sepharose for isolating ppGpp-binding proteins. A novel ppGpp-binding protein, YjgA, was isolated and characterized by MALDI-TOF MS (matrix-assisted laser desorption ionization-time-of-flight mass spectrometry) coupled with two-dimensional gel electrophoresis. YjgA and truncated forms of YjgA were cloned and over-expressed in BL21 (DE3). The binding affinity of YjgA to ppGpp was determined by equilibrium dialysis. The interaction of YjgA with ppGpp was very specific, considering that the dissociation constant of YjgA with ppGpp was measured as $5.2{\pm}2.0{\mu}M$, while the affinities to GTP and GDP were about 60 and 30 times weaker than ppGpp. Expression of yjgA gene in Escherichia coli K-12 MG1655 was examined by reverse transcription polymerase chain reaction (RT-PCR). RT-PCR results revealed that yjgA was expressed from early to late stationary phase. The yjgA deletion mutant exhibited decreased cell number at stationary phase compared to parent strain and the over-expression of YjgA increased the cell number. These results suggested that YjgA might stimulate cell division under stationary phase. In most prokaryotic genome, about half of the protein candidates are hypothetical, that are expected to be expressed but there is no experimental report on their functions. The approach utilized in this study may serve as an effective mean to probe the functions of hypothetical proteins.

• Journal of Life Science
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• v.20 no.5
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• pp.662-669
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• 2010

Lipolytic enzyme-producing bacteria were isolated from edible oil mill effluents on tributyrin agar medium. The shake-flask-scale studies yielded a promising isolate and it was identified as Pseudomonas sp. An OME using various microbiological observations such as cultural, microscopic, and biochemical tests was undertaken and confirmed using PIBWIN bacterial identification software. Lipolytic enzyme production was screened with oils such as sunflower, caster, coconut, tributyrin, and olive. Amongst these, olive oil showed an increased lipase production 6.1 U/ml. In view of the highest lipolytic enzyme production with olive oil, further optimizations were carried out using olive oil as a carbon source. Lipolytic enzyme production was optimized by a conventional 'one variable at a time' approach and the significant factors were further analyzed statistically using response surface methodology (RSM). The effect of physical factors such as incubation time, temperature, initial medium pH, and nutritional factors such as concentration of olive oil and yeast extract were examined for lipase production. Lipolytic enzyme secretion was strongly affected by three variables (incubation time, concentration of yeast extract and olive oil). Therefore, the interaction of these three factors was further optimized using response surface methodology. The optimized conditions of lipase production using response surface methodology yielded a maximum of 9.62 U/ml with optimum conditions for incubation, yeast extract and olive oil concentrations were found to be 48 hr, 0.3 g. and 0.9 ml. respectively.

• Molecules and Cells
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• v.38 no.1
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• pp.40-50
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• 2015

In the interaction between plants and pathogens, carbon (C) resources provide energy and C skeletons to maintain, among many functions, the plant immune system. However, variations in C availability on pathogen associated molecular pattern (PAMP) triggered immunity (PTI) have not been systematically examined. Here, three types of starch mutants with enhanced susceptibility to Pseudomonas syringae pv. tomato DC3000 hrcC were examined for PTI. In a dark period-dependent manner, the mutants showed compromised induction of a PTI marker, and callose accumulation in response to the bacterial PAMP flagellin, flg22. In combination with weakened PTI responses in wild type by inhibition of the TCA cycle, the experiments determined the necessity of C-derived energy in establishing PTI. Global gene expression analyses identified flg22 responsive genes displaying C supply-dependent patterns. Nutrient recycling-related genes were regulated similarly by C-limitation and flg22, indicating re-arrangements of expression programs to redirect resources that establish or strengthen PTI. Ethylene and NAC transcription factors appear to play roles in these processes. Under C-limitation, PTI appears compromised based on suppression of genes required for continued biosynthetic capacity and defenses through flg22. Our results provide a foundation for the intuitive perception of the interplay between plant nutrition status and pathogen defense.

• Journal of Life Science
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• v.31 no.1
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• pp.100-108
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• 2021

In the oral cavity, there are hundreds of microbial species that exist as planktonic cells or are incorporated into biofilms. The accumulation and proliferation of pathogenic bacteria in the oral biofilm can lead to caries and periodontitis, which are typical oral diseases. The oral bacteria in the biofilm not only can resist environmental stress inside the oral cavity, but also have a 1,000 times higher resistance to antibiotics than planktonic cells by genes exchange through the interaction between cells in the oral biofilm. Therefore, if the formation of oral biofilm is suppressed or removed, oral diseases caused by bacterial infection can be more effectively prevented or treated. In particular, since oral biofilms have the characteristic of forming a biofilm by gathering several bacteria, quorum sensing, a signaling system between cells, can be a target for controlling the oral biofilm. In addition, a method of inhibiting biofilm formation by using arginine, an alkali-producing substrate of oral bacteria, is used to convert the distribution of oral microorganisms into an environment similar to that of healthy teeth or inhibit the secretion of glucosyltransferase by S. mutans to inhibit the formation of non-soluble glucans. It can be a target to control oral biofilm. This method of inhibiting or removing the oral biofilm formation rather than inducing the death of pathogenic bacteria in the oral cavity will be a new strategy that can selectively prevent or therapeutic avenues for oral diseases including dental caries.

• Microbiology and Biotechnology Letters
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• v.49 no.1
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• pp.101-110
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• 2021

Interactions between microalgae and heterotrophic bacteria are common in natural environments. This study investigated the effect of heterotrophic bacteria on the activity of the photosynthetic eukaryotic alga Chlorella sp. MF1907 when cocultured. A total of 31 heterotrophic bacterial isolates belonging to different genera were cocultured with MF1907. Interactions of the alga with Agromyces, Rhodococcus, Sphingomonas, Hyphomicrobium, Rhizobium, and Pseudomonas were positive, while those with Burkholderia, Paraburkholderia, Micrococcus, Arthrobacter, Mycobacterium, Streptomyces, Pedobacter, Mucilaginibacter, Fictibacillus, Tumebacillus, Sphingopyxis, and Erythrobacter were negative (p < 0.05). A turnover experiment demonstrating a switch from heterotrophic to autotrophic activity of MF1907 was performed using 16 isolates exhibiting apparent effects (positive, negative, or neutral). Compared with the results of the coculture experiment, eight isolates exhibited the same outcomes, while the others did not. Consistently, Pseudomonas and Agromyces showed a remarkable positive effect on MF1907 activity, and Burkholderia, Streptomyces, and Erythrobacter had a marked negative effect. Our results suggest that it may be possible to use the isolates for controlling populations of microalgae in natural and engineered environments.

• Pediatric Infection and Vaccine
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• v.29 no.2
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• pp.70-76
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• 2022

Coronavirus disease 2019 (COVID-19) in patients with underlying diseases, is associated with high infection and mortality rates, which may result in acute respiratory distress syndrome and death. Mucopolysaccharidosis (MPS) type II is a progressive metabolic disorder that stems from cellular accumulation of the glycosaminoglycans, heparan, and dermatan sulfate. Upper and lower airway obstruction and restrictive pulmonary diseases are common complaints of patients with MPS, and respiratory infections of bacterial or viral origin could result in fatal outcomes. We report a case of COVID-19 in a 16-year-old adolescent with MPS type II, who had been treated with idursulfase since 5 years of age. Prior to infection, the patient's clinical history included developmental delays, abdominal distension, snoring, and facial dysmorphism. His primary complaints at the time of admission included rhinorrhea, cough, and sputum without fever or increased oxygen demand. His heart rate, respiratory rate, and oxygen saturation were within the normal biological reference intervals, and chest radiography revealed no signs of pneumonia. Consequently, supportive therapy and quarantine were recommended. The patient experienced an uneventful course of COVID-19 despite underlying MPS type II, which may be the result of an unfavorable host cell environment and changes in expression patterns of proteins involved in interactions with viral proteins. Moreover, elevated serum heparan sulfate in patients with MPS may compete with cell surface heparan sulfate, which is essential for successful interaction between the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein and the host cell surface, thereby protecting against intracellular penetration by SARS-CoV-2.

• Clean Technology
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• v.29 no.2
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• pp.135-144
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• 2023

Pseudomonas aeruginosa and Staphylococcus aureus are the two most frequently encountered pathogens responsible for chronic wound infections, often coexisting in such cases. These infections exhibit heightened virulence compared to single infections, leading to unfavorable patient outcomes. The interaction among microorganisms within polymicrobial infections has been shown to exacerbate disease progression. Polymicrobial infections, prevalent in various contexts such as the respiratory tract, wounds, and diabetic foot, typically involve diverse microorganisms, with Pseudomonas aeruginosa and Staphylococcus aureus being the most commonly identified pathogens. This study aimed to compare the growth patterns of bacteria under a concentration gradient of toxic chemicals, focusing on a Gram-negative strain of Pseudomonas aeruginosa and a Gram-positive strain of Staphylococcus aureus. The minimum inhibitory concentration (MIC), which signifies the concentration at which bacterial growth is inhibited, was determined by performing broth microdilution and assessing the bacteria's growth curves. The growth curves of both Pseudomonas aeruginosa and Staphylococcus aureus were confirmed, and the exponential growth phases were applied to calculate the doubling times of bacteria. The MIC value for each toxic chemical was determined through broth microdilution. These results allowed for the identification of disparities in growth rates between Gram-positive and Gram-negative bacteria, as well as differences in resistance to individual toxic substances. We expect that this approach has a strong potential for further development towards the innovative treatment of bacteria-associated infections.

• Animal Bioscience
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• v.37 no.2
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• pp.228-239
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• 2024

Objective: The objective of the experiment was to study yeast supplementation (yeast, Y) and dried kratom leaves (DKTL) on the digestibility, ruminal fermentation, blood metabolites and nitrogen balance in goats. Methods: Four of 7 to 8 months old male crossbred (50% Thai Native-Anglo Nubian) goats with average liveweight 20±0.13 kg were randomly assigned according to a 2×2 factorial arrangement in a 4×4 Latin square design to receive four diets ad libitum basis. The study investigated the effects of two levels of yeast (Y) supplementation (Y, 0 and 0.5g/kg dry matter [DM]) along with two levels of DKTL supplementation (DKTL, 0 and 4.44g/kg DM). The experimental groups were as follows: T1 = control group with 0Y+0DKTL, T2 = 0Y+4.44 DKTL, T3 = 0.5Y+0DKTL, and T4 = 0.5Y+4.44 DKTL. Results: The results showed that there were no interactions between Y levels and DKTL levels with respect to total DM intake, but there were significant effects (p<0.05) by levels of Y; goats receiving 0.05 g/kg DM Y had higher than goats fed 0.0 g/kg DM on average (kg/d). A percentage of body weight (% BW) and grams per kilogram of metallic weight (g/kg w^0.75) had no influence on yeast levels and DKTL, but there was a difference (p<0.05) by yeast level Y at 0.5 g/kg DM, being higher compared to the non-supplemented group. Apparent digestibility coefficient of nutrition in the form of (DM, organic matter, crude protein, neutral detergent fiber, and acid detergent fiber) was an increased trend in the Y-level complementary group at 0.5 g/kg DM and DKTL at 4.44 g/kg DM, respectively. Protozoa populations decreased in the group receiving Y levels at 0.5 g/kg DM and DKTL levels at 4.44 g/kg DM when compared to group T1. The acetic acid concentration and methane gas generation decreased (p<0.05) in the group receiving Y levels of 0.5 g/kg DM and DKTL levels of 4.44 g/kg DM, while the amount of propionic acid increased (p<0.05). Conclusion: Effects of feeding combinations of Y and DKTL supplementation on feed showed no interaction effect (Y×DKTL) on feed intake, rumen fermentation, bacterial and fungi population. The effect on protozoal populations was lower in the group that was supplemented with DKTL at 4.44 g/kg DM related to synthetic CH₄ was reduced.

• Journal of Life Science
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• v.30 no.9
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• pp.804-812
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• 2020

The giant mealworm beetle, Zophobas atratus (Coleoptera: Tenebrionidae) has been used as a protein source for small pets and mammals. Recently, it was temporarily registered in the list of the Food Code. We previously performed an in silico analysis of the Zophobas atratus transcriptome to identify putative antimicrobial peptides and identified several antimicrobial peptide candidates. Among them, we assessed the antimicrobial and anti-inflammatory activities of zophobacin 1 that was selected bio-informatically based on its physicochemical properties against microorganisms and mouse macrophage Raw264.7 cells. Zophobacin 1 showed antimicrobial activities against microorganisms without inducing hemolysis and decreased the nitric oxide production of the lipopolysaccharide-induced Raw264.7 cells. Moreover, ELISA and Western blot analysis revealed that zophobacin 1 reduced expression levels of pro-inflammatory enzymes such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). We also investigated expression of pro-inflammatory cytokines (interleukin-6 and interleukin-1β) production through quantitative real time-PCR and ELISA. Zophobacin 1 markedly reduced the expression level of cytokines through the regulation of mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-κB) signaling. We confirmed that zophobacin 1 bound to bacterial cell membranes via a specific interaction with lipopolysaccharides. These data suggest that zophobacin 1 could be promising molecules for development as antimicrobial and anti-inflammatory therapeutic agents.

• Korean Journal of Microbiology
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• v.47 no.3
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• pp.200-208
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• 2011

Most problematic antibiotic resistance mechanism for MLS (macrolide-lincosamide-streptogramn B) antibiotics encountered in clinical practice is mono- or dimethylation of specific adenine residue at 2058 (E. coli coordinate) of 23S rRNA which is performed by Erm (erythromycin ribosome resistance) protein through which bacterial ribosomes reduce the affinity to the antibiotics and become resistant to them. ErmSF is one of the four gene products produced by Streptomyces fradiae to be resistant to its own antibiotic, tylosin. Unlike other Erm proteins, ErmSF harbors idiosyncratic long N-terminal end region (NTER) 25% of which is comprised of arginine well known to interact with RNA. Furthermore, NTER was found to be important because when it was truncated, most of the enzyme activity was lost. Based on these facts, capability of NTER peptide to inhibit the enzymatic activity of ErmSF was sought. For this, expression system for two different proteins to be expressed in one cell was developed. In this system, two plasmids, pET23b and pACYC184 have unique replication origins to be compatible with each other in a cell. And expression system harboring promoter, ribosome binding site and transcription termination signal is identical but disparate amount of protein could be expressed according to the copy number of each vector, 15 for pACYC and 40 for pET23b. Expression of NTER peptide in pET23b together with ErmSF in pACYC 184 in E. coli successfully gave more amounts of NTER than ErmSF but no inhibitory effects were observed suggesting that there should be dynamicity in interaction between ErmSF and rRNA rather than simple and fixed binding to each other in methylation of 23S rRNA by ErmSF.