• Fisheries and Aquatic Sciences
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• v.1 no.2
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• pp.201-208
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• 1998

A protease, which had no tryptic and chymotryptic activity, was purified from the hepatopancreas of shrimp, P. japonicus, through ammonium sulfate fractionation, Q­Sepharose ionic exchange, benzamidine Sepharose 6B affinity, and Sephacryl S-100 gel chromatography. Molecular weight (M.W.) of the protease was estimated to be 24 kDa by gel filtration and showed a single peptide band by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). The protease had a low ratio of acidic to basic amino acids, which is different with pro teases from marine animals. The enzyme was partially inhibited by benzamidine, tosyl-L-lysine chioromethyl ketone (TLCK), phenylmethylsulfonyl fluoride (PMSF), soybean trypsin inhibitor (SBTI), and pepstatin. The enzyme did not have any activity against benzoyl-D,L-arginine p-nitroanilide (BAPNA) or benzoyl-L-tyrosine ethyl ester (BTEE) which is a specific substrate of trypsin and chymotrypsin, respectively. However, the enzyme showed activity forward N-CBZ-L-tyrosine p-nitrophenyl ester (CBZ-Tyr-pNE), N­CBZ-L-tryptophan p-nitrophenyl ester (CBZ-Trp-pNE), and N-CBZ-L-proline p-nitrophenyl ester (CBZ-Pro-pNE). The protease did not showed tryptic and chymotryptic activity, which was not reported in shrimp hepatopancreas.

• Bulletin of the Korean Chemical Society
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• v.28 no.6
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• pp.947-952
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• 2007

Mycobacterium tuberculosis is a pathogen responsible for 2-3 million deaths every year worldwide. The emergence of drug-resistant and multidrug-resistant tuberculosis has increased the need to identify new antituberculosis targets. Acetohydroxy acid synthase, (AHAS, EC 2.2.1.6), an enzyme involved in branched-chain amino acid synthesis, has recently been identified as a potential anti-tuberculosis target. To assist in the search for new inhibitors and “receptor-based” design of effective inhibitors of tubercular AHAS (TbAHAS), we constructed four different structural models of TbAHAS and used one of the models as a target for virtual screening of potential inhibitors. The quality of each model was assessed stereochemically by PROCHECK and found to be reliable. Up to 89% of the amino acid residues in the structural models were located in the most favored regions of the Ramachandran plot, which indicates that the conformation of each residue in the models is good. In the models, residues at the herbicide-binding site were highly conserved across 39 AHAS sequences. The binding mode of TbAHAS with a sulfonylurea herbicide was characterized by 32 hydrophobic interactions, the majority of which were contributed by residue Trp516. The model based on the highest resolution X-ray structure of yeast AHAS was used as the target for virtual screening of a chemical database containing 8300 molecules with a heterocyclic ring. We developed a short list of molecules that were predicted to bind with high scores to TbAHAS in a conformation similar to that of sulfonylurea derivatives. Five sulfonylurea herbicides that were calculated to efficiently bind TbAHAS were experimentally verified and found to inhibit enzyme activity at micromolar concentrations. The data suggest that this time-saving and costeffective computational approach can be used to discover new TbAHAS inhibitors. The list of chemicals studied in this work is supplied to facilitate independent experimental verification of the computational approach.

• Journal of Microbiology and Biotechnology
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• v.29 no.4
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• pp.587-595
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• 2019

Pharmacological research on (CHA), a marine-derived quinazolinone alkaloid with significant cytotoxic activity, is restricted by low yields and is a problem that needs to be settled urgently. In this work, the selection of additional nitrogen sources and the optimization of additional concentrations and longer fermentation times using ammonium acetate, were investigated. CHA production was optimized to 62.1 mg/l with the addition of 50 mM ammonium acetate at 120 h of the fermentation in the shaker flask. This feeding strategy significantly increased 3-deoxy-arabino-heptulosonate-7-phosphate synthase activity and transcript levels of critical genes (laeA, dahp, and trpC) in the shikimate pathway compared with the non-treatment group. In addition, the selection of the feeding rate (0.01 and $0.03g/l{\cdot}h$) was investigated in a 5-L bioreactor. As a result, CHA production was increased by 57.9 mg/l with a $0.01g/l{\cdot}h$ ammonium acetate feeding rate. This work shows that the strategy of ammonium acetate supplementation had an effective role in improving CHA production by Aspergillus fumigatus CY018. It also shows that this strategy could serve as an important example of large-scale fermentation of a marine fungus in submerged culture.

• Asian-Australasian Journal of Animal Sciences
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• v.32 no.3
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• pp.430-436
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• 2019

Objective: This study identified angiotensin I-converting enzyme (ACE) inhibitory peptides in beef M. longissimus injected with thermolysin (80 ppm) and stored for 3 days at $5^{\circ}C$. Methods: Crude peptides (molecular weight <3 kDa) were obtained from the thermolysin hydrolysate and separated into seven fractions. Fraction V showing the highest ACE inhibitory activity was further fractionated, yielding subfractions V-15, V-m1, and V-m2, and selected for superior ACE inhibitory activity. Finally, twelve peptides were identified from the three peak fractions and the ACE inhibitory activity ($IC_{50}$) of each peptide was evaluated. Results: The Leu-Ser-Trp, Phe-Gly-Tyr, and Tyr-Arg-Gln peptides exhibited the strongest ACE inhibitory activity ($IC_{50}$ values of 0.89, 2.69, and 3.09 mM, respectively) and had higher concentrations (6.63, 10.60, and 29.91 pg/g; p<0.05) relative to the other peptides tested. Conclusion: These results suggest that the thermolysin injection process is beneficial to the generation of bioactive peptides with strong ACE inhibitory activity.

• Journal of Microbiology and Biotechnology
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• v.31 no.1
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• pp.25-32
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• 2021

Inflammatory reactions activated by lipopolysaccharide (LPS) of gram-negative bacteria can lead to severe septic shock. With the recent emergence of multidrug-resistant gram-negative bacteria and a lack of efficient ways to treat resulting infections, there is a need to develop novel anti-endotoxin agents. Antimicrobial peptides have been noticed as potential therapeutic molecules for bacterial infection and as candidates for new antibiotic drugs. We previously designed the 9-meric antimicrobial peptide Pro9-3 and it showed high antimicrobial activity against gram-negative bacteria. Here, to further examine its potency as an anti-endotoxin agent, we examined the anti-endotoxin activities of Pro9-3 and elucidated its mechanism of action. We performed a dye-leakage experiment and BODIPY-TR cadaverine and limulus amebocyte lysate assays for Pro9-3 as well as its lysine-substituted analogue and their enantiomers. The results confirmed that Pro9-3 targets the bacterial membrane and the arginine residues play key roles in its antimicrobial activity. Pro9-3 showed excellent LPS-neutralizing activity and LPS-binding properties, which were superior to those of other peptides. Saturation transfer difference-nuclear magnetic resonance experiments to explore the interaction between LPS and Pro9-3 revealed that Trp3 and Tlr7 in Pro9-3 are critical for attracting Pro9-3 to the LPS in the gram-negative bacterial membrane. Moreover, the anti-septic effect of Pro9-3 in vivo was investigated using an LPS-induced endotoxemia mouse model, demonstrating its dual activities: antibacterial activity against gram-negative bacteria and immunosuppressive effect preventing LPS-induced endotoxemia. Collectively, these results confirmed the therapeutic potential of Pro9-3 against infection of gram-negative bacteria.

• Animal Bioscience
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• v.36 no.10
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• pp.1584-1595
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• 2023

Objective: This study was conducted to evaluate the effect of a low-protein diet on growth performance, carcass traits, nutrient digestibility, blood profiles, and odor emissions in growing-finishing pigs. Methods: A total of 126 crossbred pigs ([Yorkshire×Landrace]×Duroc) with an average body weight (BW) of 38.56±0.53 kg were used for a 14-week feeding trial. Experimental pigs were allotted to one of 6 treatments in 3 replicates of 7 pigs per pen in a randomized complete block design. Pigs were fed each treatment diet with different levels of crude protein (CP). Phase 1 (early growing): 14%, 15%, 16%, 17%, 18%, 19%; phase 2 (late growing): 13%, 14%, 15%, 16%, 17%, 18%; phase 3 (early finishing): 12%, 13%, 14%, 15%, 16%, 17%; phase 4 (late finishing): 11%, 12%, 13%, 14%, 15%, 16%. All experimental diets in each phase were contained the same concentration of lysine (Lys), methionine (Met), threonine (Thr), and tryptophan (Trp). Results: Over the entire experimental period, there was no significant difference in BW, average daily feed intake, and gain-to-feed ratio among all treatments (p>0.05), but a quadratic effect (p = 0.04) was observed in average daily gain (ADG) during the late finishing phase with higher ADG in Group D. Blood urea nitrogen concentration linearly increased with an increase in dietary CP levels (p<0.01). Regarding nutrient digestibility, excreted nitrogen in urine and feces and nitrogen retention linearly increased as the CP level increased (p<0.01). A linear effect was observed with increasing CP levels in amines, ammonia, and hydrogen sulfide in odor emissions (p<0.01). No significant effects were observed in the measurements of carcass traits and meat characteristics (p>0.05). Conclusion: In phase feeding, reducing the CP level to 14% in early-growing pigs, 13% in late-growing pigs, 12% in early-finishing pigs, and 11% in late-finishing pigs is recommended.

• Journal of The Korean Society of Inherited Metabolic disease
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• v.16 no.2
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• pp.93-101
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• 2016

Purpose: McArdle disease, glycogen storage disease type V (GSD V), is one of the most common adolescent-onset glycogen storage diseases. It is caused by recessive mutations in PYGM encoding myophosphorylase, which is critical to glycogen metabolism. Since only a few korean patients have been reported, we will observe the clinical and genetic features of three korean patients with McArdle disease. Methods: We retrospectively reviewed the medical records of three patients with genetically confirmed McArdle disease, including the results of forearm ischemic exercise test, electromyogram, nerve conduction velocity, muscle biopsy, and PYGM analysis in peripheral leukocytes. Results: All three cases were males and their age of symptom onset was 12, 5, 14 years old, respectively. A high basal level of serum creatine kinase was noted in all three patients. They experienced the recurrent episodes of rhabdomyolysis, but second wind phenomenon was not definite. In muscle biopsy, subsarcolemmal space vacuoles including periodic acid schiff stained materials were found in two patients, while no evidence of glycogen storage disease was found in the other. A total of five different mutations, $p.Arg50^*$, p.Trp798Arg, $p.Arg50^*$, p.Glu779del, $p.Asp511Thrfs^*28$ and p.Phe710del, were found in three patients. Avoidance of isometric exercise, aerobic exercise and glucose intake before each exercise were recommended for all patients. Conclusion: The three Korean patients with McArdle disease showed the typical manifestations of the condition. The most mutations were private. Therefore, identification of more cases with long-term follow-up will be required to understand the clinical and genetic features of this disease among Korean population.

• Journal of the Korean Society of Food Science and Nutrition
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• v.32 no.2
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• pp.211-216
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• 2003

This study was carried out to determine the antioxidative, antimutagenic, and anticancer effects of Rhodiola sachalinensis root using 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical donating method, Ames test and cytotoxicity, respectively. Rhodiola sachalinenis root were extracted with ethanol and then further fractionated to n-hexane, chloroform, ethyl acetate (EtOAc), butanol and water, stepwise. Among five fractions, the Etohc fractions showed the highest electron donating activities (14.3 $\mu$g/mL). The inhibition rate of ethanol extract (200$\mu$g/plate) of Rhodiola sachalinensis root in the S. typhimurium TA100 strain showed 89.1% inhibition against the mutagenesis induced by MNNG. In addition, the suppression of EtOAc fractions with same concentration of Rhodiola sachalinensis root in the S. typhimurium TA98 and TAI00 strains showed 89.7% and 91.5% inhibition against 4NQO, respectively. The suppressions under the same condition against B($\alpha$)P and Trp-P-1 in the TA98 and TA100 strains were 94.2% and 95.7%, and 92.3% and 93.8%, respectively. The cytotoxic effects of Rhodiola sachalinensis root against the cell lines with human lung carcinoma (A549), human hepatocellular carcinoma (HepG2), human gastric carcinoma (AGS) and human breast adenocarcinoma (MCF-7) were inhibited with the increase of the extract concentration. The treatment of 1.0 mg/mL Rhodiola sachalinensis root of EtOAc fraction showed strong cytotoxicities of 90.5%, 81.5%, 92.2% and 82.6% against A549, HepG2, AGS and MCF-7, respectively.

• Journal of The Korean Society of Inherited Metabolic disease
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• v.18 no.1
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• pp.30-34
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• 2018

Niemann Pick type C disease (NPC) is an inherited progressive neurodegenerative disorder, due to defects of intracellular lipid trafficking and storage. Hepatosplenomegaly may prevail, while progressive neurodegenerative symptoms such as cerebellar involvement, dystonia, vertical supranuclear ophthalmoplegia, cataplexy, and eventually seizures starting at juvenile or late infantile period may accompany after normal early development. Here we describe a 3-year-old Korean boy with NPC who presented with splenomegaly at age 3. Liver biopsy showed characteristic foamy cell stained by periodic acid-schiff, and molecular analysis for NPC1 identified the compound heterozygous mutations, novel mutation of c.1631G>A (p.Trp544Ter) and c.2662C>T (p. Pro888Ser) as a known mutation. Filipin was strongly stained with unesterified cellular cholesterol in the patient's skin fibroblasts. The patient has received migulstat since age 3 years and his long-term outcome is needed to be observed.

• Journal of the Korean Society of Food Science and Nutrition
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• v.31 no.2
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• pp.322-328
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• 2002

This study was carried out to investigate antimutagenic and anticancer effects of ethanolic extract of Korean traditional doenjang added sea tangle. Most of the mineral content of doeniang was increased by addition of sea tangle. In the Ames test, the antimutagenic effect of ethanol extract of Korean traditional doenjang added 5% sea tangle was higher than that of control (no additive), 10%, and 15% sea tangle additions. The inhibition rate of ethanol extract (200$\mu\textrm{g}$/plate) of doenjang added 5% sea tangle in the S. typhimurium TA100 strain showed 97.0% inhibition against the mutagenesis induced by MNNG. In addition, the suppression of ethanol extract (200$\mu\textrm{g}$/plate) of doenjang added 5% sea tangle in the S. typhimurium TA98 and TA100 strains showed 60.2% and 69.1% inhibition respectively, against the mutagenesis induced by 4NQO. The suppressions under the same condition against B($\alpha$ )P and Trp-P-1 in the TA98 and TA100 strains were 71.7% and 87.3%, and 66.6% and 80.8%, respectively. In the anticancer effects, the cytotoxicity of doenjang added 5% sea tangle on the cell lines with human lung carcinoma (A549), human hepatocellular carcinoma (HepG2), and human gastric carcinoma (KATOIII) were inhibited with increasing the extract concentration. The treatment of 1.0 mg/mL Doenjang added 5% sea tangle showed strong cytotoxicity of 56.4%, 87.67%, and 89.5% against A549, HepG2, and KATOIII, respectively.