• KSBB Journal
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• v.23 no.6
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• pp.474-478
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• 2008

UDP-glucose regeneration system using metabolic engineeringis unique and efficient strategy for oligosaccharide synthesis. To exploit the efficient UDP-glucose regeneration system, we introduced four enzymes, which would be important in partitioning the flux of UDP regenerationsuch as UDP-glucose pyrophosphorylase, UDP-Kinase gene, UDP-galactose 4-epimerase, and $\beta$-1, 4-galactasyltrasnsferase, into E. coli AD202. To determine the optimal expression level for UDP-regeneration, LacNAc concentration was compared depending on IPTG concentration. 0.5 mM IPTG induction showed the higher oligosaccharides synthesis. Using metabolic engineering under optimal IPTG induction, LacNAc synthesis of AD202/pQNGLU increased until 16 h and showed the 1.34 mM. This concentration is 10 times higher than that of control strain at same reaction time. Lactose of AD202/pQNGLU showed the maximum synthesis of 0.39 mM at 16 h and showed the 2.6 times higher than that of control strain.

• Journal of Environmental Analysis, Health and Toxicology
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• v.21 no.4
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• pp.243-251
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• 2018

Pharmaceuticals in wastewater effluents have been recognized as emerging pollutants threatening freshwater organisms. To extend understanding for bioaccumulation and toxicity in those organisms, information on biotransformation products (or metabolites) and their metabolic pathway are crucial. The aim of the present study is to identify and elucidate metabolites of pharmaceuticals formed in exposed organisms using suspect and nontarget screening approach using LC-HRMS. As the target pharmaceuticals, carbamazepine, ketoprofen, metoprolol, propranolol, and verapamil were selected whereas Daphnia magna and Gammarus pulex were used as test organisms. After 24h exposure, metabolites formed in the organisms were identified using LC-HRMS. The structures of metabolites were elucidated via analysis of MS/MS fragment pattern and the comparison with fragment database. As the results, a total of 10 metabolites were identified for 5 parent compounds (C253/C356 for carbamazepine, K211 for ketoprofen, M256 for metoprolol, P218/P276/P306 for propranolol, V196/V291/V441 for verapamil). Among them, the presence of C253 and V291 was confirmed using standard materials. Most of the identified metabolites were formed through oxidative reactions such as hydroxylation, N-demethylation, and dealkylation. Cysteine conjugation (phase II reaction) metabolite (C356) for carbamazepine was found in daphnia. The metabolic pathway of verapamil showed similar metabolic pathways and metabolic pathways for both species. Although the toxicological information on the identified metabolites could not be confirmed, the molecular structure information of the proposed metabolites can be used for future evaluation and prediction of toxicity.

• Nutrition Research and Practice
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• v.15 no.2
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• pp.160-172
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• 2021

BACKGROUND/OBJECTIVES: Nutritional status and food intake during pregnancy and lactation can affect fetal programming. In the current metabolic syndrome epidemic, high-fructose diets have been strongly implicated. This study investigated the effect of maternal high-fructose intake during pregnancy and lactation on the development of metabolic syndrome in adult offspring. SUBJECTS/METHODS: Drinking water with or without 20% fructose was administered to female C57BL/6J mice over the course of their pregnancy and lactation periods. After weaning, pups ate regular chow. Accu-Chek Performa was used to measure glucose levels, and a tail-cuff method was used to examine systolic blood pressure. Animals were sacrificed at 7 months, their livers were excised, and sections were stained with Oil Red O and hematoxylin and eosin (H&E) staining. Kidneys were collected for gene expression analysis using quantitative real-time Polymerase chain reaction. RESULTS: Adult offspring exposed to maternal high-fructose intake during pregnancy and lactation presented with heavier body weights, fattier livers, and broader areas under the curve in glucose tolerance test values than control offspring. Serum levels of alanine aminotransferase, aspartate aminotransferase, glucose, triglycerides, and total cholesterol and systolic blood pressure in the maternal high-fructose group were higher than that in controls. However, there were no significant differences in mRNA expressions of renin-angiotensin-aldosterone system genes and sodium transporter genes. CONCLUSIONS: These results suggest that maternal high-fructose intake during pregnancy and lactation induces metabolic syndrome with hyperglycemia, hypertension, and dyslipidemia in adult offspring.

• Journal of Microbiology and Biotechnology
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• v.33 no.10
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• pp.1351-1360
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• 2023

Endocrine-disrupting chemicals (EDCs) are compounds that disturb hormonal homeostasis by binding to receptors. EDCs are metabolized through hepatic enzymes, causing altered transcriptional activities of hormone receptors, and thus necessitating the exploration of the potential endocrine-disrupting activities of EDC-derived metabolites. Accordingly, we have developed an integrative workflow for evaluating the post-metabolic activity of potential hazardous compounds. The system facilitates the identification of metabolites that exert hormonal disruption through the integrative application of an MS/MS similarity network and predictive biotransformation based on known hepatic enzymatic reactions. As proof-of-concept, the transcriptional activities of 13 chemicals were evaluated by applying the in vitro metabolic module (S9 fraction). Identified among the tested chemicals were three thyroid hormone receptor (THR) agonistic compounds that showed increased transcriptional activities after phase I+II reactions (T3, 309.1 ± 17.3%; DITPA, 30.7 ± 1.8%; GC-1, 160.6 ± 8.6% to the corresponding parents). The metabolic profiles of these three compounds showed common biotransformation patterns, particularly in the phase II reactions (glucuronide conjugation, sulfation, GSH conjugation, and amino acid conjugation). Data-dependent exploration based on molecular network analysis of T3 profiles revealed that lipids and lipid-like molecules were the most enriched biotransformants. The subsequent subnetwork analysis proposed 14 additional features, including T4 in addition to 9 metabolized compounds that were annotated by prediction system based on possible hepatic enzymatic reaction. The other 10 THR agonistic negative compounds showed unique biotransformation patterns according to structural commonality, which corresponded to previous in vivo studies. Our evaluation system demonstrated highly predictive and accurate performance in determining the potential thyroid-disrupting activity of EDC-derived metabolites and for proposing novel biotransformants.

• Journal of Life Science
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• v.16 no.2 s.75
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• pp.199-205
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• 2006

Peroxisome proliferator activated receptor (PPAR)-$\alpha$ of three PPAR subtypes ($-\alpha,\;-\beta/-\gamma,\;-\delta$), which are members of the nuclear hormone receptor superfamily of ligand-activated transcription factors, plays a key role in lipoprotein and glucose homeostasis. A variation in the PPAR-a gene expression has been suggested to influence the development of metabolic syndrome through alterations in lipid concentrations. The aim of our study was to investigate the association between the PPAR-a and metabolic syndrome among South Korean. A total of 542 health screen examinees were enrolled in this study who were examined in Kosin University Gospel Hospital from December, 2004 to July, 2005. The height, weight, waist circumference, and systolic and diastolic blood pressure of the subjects were examined and fasting blood glucose, total cholesterol, HDL cholesterol, LDL cholesterol, triglyceride were measured by-sampling in venous blood. The metabolic syndrome was defined as the presence of three or more of the following : waist circumference men ${\geq}90cm$, women ${\geq}80cm$, blood pressure ${\geq}130/85mmHg$, fasting glucose ${\geq}110mg/dL$, HDL cholesterol men <40 mg/dL, women <50 mg/dL, triglyceride ${\geq}150mg/dL$. The blood pressure, fasting glucose, HDL cholesterol, triglyceride were evaluated by using the criteria of NECP ATP III and waist circumference was assessed by using the criteria of WHO Asia-Western Pacific. And the author compared the frequency of the PPAR-$\alpha$ mutation of L162V ($C{\rightarrow}G$ variant in exon 5) in a sample of 542 subjects with and without the metabolic syndrome by polymerase chain reaction allele-specific oligonucleotide (PCR-ASO) method. One (0.2%) hetero-isotype among high risk of metabolic syndrome was identified. The values of waist circumference, body mass index and low density lipoprotein cholesterol of the mutant were 100 cm, 28.6 $kg/m^2$ and 120 mg/dL, respectively. Although the author failed to see significant association between the presence of the PPAR-$\alpha$ L162V polymorphism and metabolic syndrome, one PPAR-$\alpha$ L162V polymorphism in metabolic syndrome patients was found.

• Journal of Microbiology and Biotechnology
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• v.7 no.3
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• pp.212-214
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• 1997

A new isolate, Bacillus sp. ALK-7 can synthesize ethylene from l-aminocyclopropane-l-carboxylic acid (ACC) as well as from methionine. The ACC has only been recognized as a key intermediate found in the metabolic pathway leading to ethylene formation in various plants. The efficiency of ethylene formation from the ACC by Bacillus sp. ALK-7 was about 2 times as high as that from the methionine. The reaction from ACC to ethylene formation was also shown to be mediated by the cell-free extracts of Bacillus sp. ALK-7.

• Korean Journal of Veterinary Service
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• v.36 no.4
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• pp.311-320
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• 2013

The purpose of this study was to evaluate detecting methods and the relationship between tissues and blood for enrofloxacin and metabolic ciprofloxacin residues in broiler chickens. Two groups of broiler chickens were administrated via the drinking water with $50{\mu}g/mL$ and $100{\mu}g/mL$ of enrofloxacin for 5 days, respectively. The concentration of enrofloxacin and metabolic ciprofloxacin in tissues (muscle and kidney) and blood were measured during administration period (for 5 days) and withdrawal period (for 12 days) by high performance liquid chromatography (HPLC) method. Also, all samples were conducted for screening of residues by microbial method using E. coli for quinolone detection and immuno-chromatography method using Smart kit. The relationship between tissues (muscle and kidney) and blood for enrofloxacin and metabolic ciprofloxacin residues in broiler chickens was followed : The levels of enrofloxacin and metabolic ciprofloxacin residues in muscle and kidney were higher 2.9~3.2 folds, 3.6~3.8 folds more than the residues levels in blood, respectively. These results support we can predict the residues in muscle and kidney from the residues in blood. In comparison of detecting methods for antibiotic residues, microbial method using E. coli for quinolone detection and immuno-chromatography method using Smart kit could detect positive reaction at similar or lower concentration than violative concentration of enrofloxacin and metabolic ciprofloxacin in chicken tissues. These results support what two screening methods are useful for screening of quinolone detection in chickens.

• Proceedings of the Korean Society of Soil and Groundwater Environment Conference
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• 1998.06a
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• pp.53-60
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• 1998

The effect of chemically oxidized intermediated of PAH compounds on the degradation of the parent PAHs was characterized and evaluated for the context of cooxidation. Anthracene and pyrene exhibited extensive degradation (mean percent removal of 57.5%) after 28 days of incubation by introducing the Fenton oxidation intermediate of the PAH compounds, while unoxidized anthracene and pyrene exhibited 12.5% removal. Dehydrogenase activities for the oxidized PAH studies ware enhanced two to five folds to the unoxidized PAHs studies. The chemical oxidation products can serve as a structually very similar analogue substrates for a consortia of soil microorganisms and as a metabolic intermediates in the biodegradation sequence of the parent PAH compounds. These results may be interpreted in the context of cooxidation mechanism whereby high recalcitrant PAH compounds are biodegraded in the soil and suggest a potential tool for bioremediation of PAHs contaminated soils and protection of groundwater.

• Proceedings of the Korean Society of Life Science Conference
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• 2001.11a
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• pp.3-8
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• 2001

Isopentenyl diphosphate (IPP) is the common, five-carbon building block in the biosynthesis of all carotenoids, IPP in Escherichia coli is synthesized through the non-mevalonate pathway. The first reaction of IPP biosynthesis in E. coli is the formation of 1-deoxy-D-xylulose-5-phosphate (DXP), catalyzed by DXP synthase and encoded by dxs. The second reaction in the pathway is the reduction of DXP to 2-C-methyl-D-erythritol-4-phosphate, catalyzed by DXP reductoisomerase and encoded by dxr. To determine if one or more of the reactions in the non-mevalonate pathway controlled flux to IPP, dxs and dxr were placed on several expression vectors under the control of three different promoters and transformed into three E. coli strains (DH5(, XL1-Blue, and JM101) that had been engineered to produce lycopene. Lycopene production was improved significantly in strains transformed with the dxs expression vectors. When the dxs gene was expressed from the arabinose-inducible araBAD promoter (PBAD) on a medium-copy plasmid, lycopene production was 2-fold higher than when dxs was expressed from the IPTG-inducible trc and lac promoters (Ptrc and Plac, respectively) on medium-copy and high-copy plasmids, Given the low final densities of cells expressing dxs from IPTG-inducible promoters, the low lycopene production was probably due to the metabolic burden of plasmid maintenance and an excessive drain of central metabolic intermediates. At arabinose concentrations between 0 and 1.33 mM, cells expressing both dxs and dxr from PBAD on a medium-copy plasmid produced 1.4 - 2.0 times more lycopene than cells expressing dxs only. However, at higher arabinose concentrations lycopene production in cells expressing both dxs and dxr was lower than in cells expressing dxs only. A comparison of the three E. coli strains transformed with the arabinose-inducible dxs on a medium-copy plamid revealed that lycopene production was highest in XL1-Blue.

• Journal of Microbiology and Biotechnology
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• v.20 no.8
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• pp.1196-1203
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• 2010

In the present work, methanethiol and dimethyldisulfide were investigated as sulfur sources for methionine synthesis in Corynebacterium glutamicum. In silico pathway analysis predicted a high methionine yield for these reduced compounds, provided that they could be utilized. Wild-type cells were able to grow on both methanethiol and dimethyldisulfide as sole sulfur sources. Isotope labeling studies with mutant strains, exhibiting targeted modification of methionine biosynthesis, gave detailed insight into the underlying pathways involved in the assimilation of methanethiol and dimethyldisulfide. Both sulfur compounds are incorporated as an entire molecule, adding the terminal S-$CH_3$ group to O-acetylhomoserine. In this reaction, methionine is directly formed. MetY (O-acetylhomoserine sulfhydrylase) was identified as the enzyme catalyzing the reaction. The deletion of metY resulted in methionine auxotrophic strains grown on methanethiol or dimethyldisulfide as sole sulfur sources. Plasmid-based overexpression of metY in the ${\Delta}$metY background restored the capacity to grow on methanethiol or dimethyldisulfide as sole sulfur sources. In vitro studies with the C. glutamicum wild type revealed a relatively low activity of MetY for methanethiol (63 mU/mg) and dimethyldisulfide (61 mU/mg). Overexpression of metY increased the in vitro activity to 1,780 mU/mg and was beneficial for methionine production, since the intracellular methionine pool was increased 2-fold in the engineered strain. This positive effect was limited by a depletion of the metY substrate O-acetylhomoserine, suggesting a need for further metabolic engineering targets towards competitive production strains.