• Bulletin of the Korean Chemical Society
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• 제25권12호
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• pp.1791-1800
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• 2004

Angiotensin I, a model decapeptide, was glycosylated and partially hydrolyzed with HCl (6 N, 80 $^{\circ}C$, 4 h), aminopeptidase, and carboxypeptidase Y. A single peptide mass map obtained from truncated peptides in the partial acid hydrolysate of angiotensin and its glycosylation product mixture by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry enabled sequencing of angiotensin by a combinatorial procedure. MALDI-TOF and electrospray ionization (ESI) tandem mass spectrometric results indicate that both the N-terminal amino group of aspartic acid and the guanidinium group of the second residue arginine are glycosylated.

• Toxicological Research
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• 제12권1호
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• pp.47-52
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• 1996

The renal toxicity of the extract of Polygalae Radix was investigated in rats. Rats were treated with 3.5 mg/Kg of the extract, i.p., for 7 days. Changes in consumatory behavior, 24 hour-urine and the activities of urinary enzymes were determined during the administration of the extract. Significant decrease in body weight and food consumption and increase in 24 hour-urine volume were observed during the administration. However, the quantity of total creatinine in urine was decreased significantly. Those indicate that subacute treatment with the extract might induce diuresis and the ditiresis might be due to the decrease in water reabsorption. In the activities of urinary enzymes, the activities of alanine aminopejotidase (AAP) and gamma-glutamyl transpeptidase (GGT) were increased 4.3 and 3.5 times and then returned to the control. The activity of N-acetyl-${\beta}$-D-glucosaminidase (NAG) was increased 7.2 times and then decreased slowly. But, it was significantly higher than that of the control evea after the last administration. The activity of factate dehydrogenase (LDH) was increased continuozlsly during the treatment. It showed 32 times higher than the control. These results suggested that the extract of Polygalae Radix had toxic effect on kidney. Furthermore, the result suggested that the subacute administration of the extract induced resistance against the toxicity of Polygalae Radix.

• Journal of Acupuncture Research
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• 제20권5호
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• pp.50-62
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• 2003

Objective: Bee Venom(BV) has been used for various kinds of inflammatory or painful conditions in Oriental Medicine clinics, and there publishes reports on its therapeutic effects and the probable mechanism of those therapeutic effects, where CDs and cytokines plays important role. This study investigated the influences of bee venom on the expressions of CDs and cytokines of HMC cell line Methods: In this study we analysed the expression profile of HMC cell line treated with BV of 10-2ug/ml in relation to that of HMC cell line treated with vehicle by way of CD/cytokine microarray hybridization with 342 genes on it. Results: There were no upregulated genes by more than 3 fold, while there showed some downregulated genes by less than 1/3 fold as follows: colony stimulating factor 2, CD122, IL-7, CD112, TNF-alpha, CD138, CD166, TGFbetaR2, CD42b, CD62L, CD111, interleukin 10 receptor alpha, colony stimulating factor 1(macrophage), CD38 antigen(p45), CD121a, CD33 antigen(gp67), colony stimulating factor 1 receptor, B cell linker protein (SLP65) mRNA, CD94, alanyl(membrane) aminopeptidase, immunoglobulin(CD79A) binding protein 1, CD205, CD241, CD207, CDw121b, integrin alpha L(CD11a), integrin beta 1(CD29), CD91, CD42b. Conclusions: Bee venom treatment induced downregulation of some CDs or cytokines including $TNF-{\alpha}$. IL-1R with its possible implication in an antiinflammatory action of BV. Further research on expression profile changes induced by BV treatment is expected.

• BMB Reports
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• 제32권3호
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• pp.239-246
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• 1999

A tripeptidase (PepT) was purified to homogeneity from Bacillus subtilis through four sequential chromatographies including DEAE-Sepharose ion exchange, hydroxylapatite, mono-Q FPLC ion exchange, and Superose-12 FPLC gel filtration. The apparent molecular mass of the enzyme was 49,200 Da and 51,400 Da as determined by sodium dodecylsulfatepolyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography, respectively, and the enzyme exists in a monomeric form. The physicochemical properties of the enzyme were as follows: optimum pH at 7.5, optimum temperature at $60^{\circ}C$, and pI at 4.9. The $K_m$ and $V_{max}$ values of the enzyme were 4.3 mM and 2.5 mmol/min/mg, respectively, with MetAla-Ser as substrate. The B. subtilis PepT requires $Co^{2+}$ ion(s) for activation, while it is inactivated by EOTA and 1,10-phenanthroline, suggesting that it is a metalloprotein. The enzyme was not inhibited by any of serine protease, aspartic protease, or leucine aminopeptidase inhibitors. The enzyme showed comparable activities towards four different substrates including Met-Ala-Ser, Leu-Gly-Gly, Leu-Ser-Phe, and Leu-Leu-Tyr. The amino terminal sequence of PepT determined by Edman degradation was found to be MKEEIIERFTTYVXV and turned out to be identical to that of PepT deduced from a cloned B. subtilis pepT.

• Journal of Ecology and Environment
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• 제33권3호
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• pp.261-270
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• 2010

Increasing atmospheric $CO_2$ affects the soil carbon cycle by influencing microbial activity and the carbon pool. In this study, the effects of elevated $CO_2$ on extracellular enzyme activities (EEA; ${\beta}$-glucosidase, N-acetylglucosaminidase, aminopeptidase) in salt marsh sediment vegetated with Suaeda japonica were assessed under ambient atmospheric $CO_2$ concentration (380 ppm) or elevated $CO_2$ concentration (760 ppm) conditions. Additionally, the community structure of sulfate-reducing bacteria (SRB) was analyzed via terminal restriction fragments length polymorphism (T-RFLP). Sediment with S. japonica samples were collected from the Hwangsando intertidal flat in May 2005, and placed in small pots (diameter 6 cm, height 10 cm). The pots were incubated for 60 days in a growth chamber under two different $CO_2$ concentration conditions. Sediment samples for all measurements were subdivided into two parts: surface (0-2 cm) and rhizome (4-6 cm) soils. No significant differences were detected in EEA with different $CO_2$ treatments in the surface and rhizome soils. However, the ratio of ${\beta}$-glucosidase activity to N-acetylglucosaminidase activity in rhizome soil was significantly lower (P < 0.01) at 760 ppm $CO_2$ than at 380 ppm $CO_2$, thereby suggesting that the contribution of fungi to the decomposition of soil organic matter might in some cases prove larger than that of bacteria. Community structures of SRB were separated according to different $CO_2$ treatments, suggesting that elevated $CO_2$ may affect the carbon and sulfur cycle in salt marshes.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1996년도 춘계학술대회
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• pp.213-213
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• 1996

The effects of the anti-tumor agent, SDZ-y2434, on rat kidney were investigated to predict the toxicities of its derivatives and to develope less toxic derivatives. After adjusted in metabolic cages for 5 days, rats were treated SDZ-62434(acute : 25mg/kg, i.p, once and 50mg/kg, i.p., once; subacute ; 10mg/kg, i.p., daily for 7 days). Kidney weights and urine volume during the treatment were observed. Creatinine concentration, protein concentration and the activities of N-acetyl-${\beta}$-D-glucosaminidase (NAG), alanine aminopeptidase (AAP), ${\gamma}$-glutamyl transpeptidase (GGT) and lactate dehydrogenase(LDH) in 24 hr urine were also determined. The kidney weights after the acute and subacute administration didn't show any difference. Urine volume increased 5 days after the acute administration (50mg/kg) and 3 days after the subacute administration. The excretion of creatinine was increased 5 days after the acute (50mg/kg) and subacute administration. However, the protein excretion didn't show any change. NAG acivity declined 7 days after the subacute administration. AAP and GGT activites increased 3 days after the acute administration (50mg/kg) but, returned to the control value. LDH activity showed continuousely high value after the subacute administration. These results indicates that the acute administration of SDZ-62434 might damage on glomerulus and that the subacute administration might be cytotoxic to kidney cells.

• Journal of Microbiology and Biotechnology
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• 제17권6호
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• pp.952-959
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• 2007

His-His-Leu (HHL), a tripeptide derived from a Korean soybean paste, is an angiotensin-I-converting enzyme (ACE) inhibitor. We report here a method of producing this tripeptide efficiently by expressing tandem multimers of the codons encoding the peptide in E. coli and purifying the HHL after hydrolysis of the peptide multimers. The HHL gene, tandemly multimerized to a 40-mer, was ligated with ubiquitin as a fusion gene (UH40). UH40 was inserted into vector pET29b; the UH40 fusion protein was then produced in E. coli BL21. The recombinant UH40 protein was purified by cation-exchange chromatography with a yield of 17.3mg/l and analyzed by matrixassisted laser desorption ionization (MALDI) time-of-flight (TOF) mass spectrometry and protein N-terminal sequencing. Leucine aminopeptidase was used to cleave a 405-Da HHL monomer from the UH40 fusion protein and the peptide was purified using reverse-phase high-performance liquid chromatography (HPLC) on a C18 HPLC column, with a final yield of 6.2mg/l. The resulting peptide was confirmed to be HHL with the aid of MALDI-TOF mass spectrometry, glutamine-TOF mass spectrometry, N-terminal sequencing, and measurement of ACE inhibiting activity. These results suggest that our production method is useful for obtaining a large quantity of recombinant HHL for functional antihypertensive peptide studies.

• 한국식품영양학회지
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• 제22권1호
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• pp.20-28
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• 2009

According to traditional oriental medicine, only non-coagulated native deer blood is said to be effective, and coagulated deer blood is ineffective. Thus, a drying and tablet-producing method for deer blood was developed to maintain its physiological and therapeutic activity, and so that after drying, it can be redissolved and protected from coagulation. Proteases such as trypsin, pepsin, chymotrypsin, and aminopeptidase were added to the deer blood indicating that it coagulated in an hour, as shown by the reference. Wax gourd extract, which is high in protease, was added to the blood resulting in anticoagulation for 31 hours. Also, additions of 1% EDTA, 0.38% sodium citrate, 0.16% calcium oxalate, 1.2% ethanol, and 0.006% heparin to the deer blood resulted in anticoagulation for 1 hour, 4 hours, 2 hours, 1 hour, and 31 hours, respectively. In an experiment using 0.19% sodium citrate plus 1% wax gourd extract, and 0.006% heparin plus 1% wax gourd extract, anticoagulation was maintained for up to 72 hours. However, since heparin can not be used in food, the deer blood tablet was made with the addition of 0.19% sodium citrate and 1% wax gourd extract, followed by freeze drying. The dissolution rate for the tablet manufactured in this manner was 96.7%. And the dissolution rates for spray-dried deer blood, vacuum-dried deer blood, and marketed deer blood tablets were 85%, 81%, and 25.5%, respectively. The composition of the tablet produced from the freeze-dried deer blood was 56.4% protein, 18.7% lactose, 1.2% amino acids, 1.0% glucose, 0.7% lipids, 180 mg/100 g of iron, 13 mg/100 g of potassium, 39.1 mg/100 g of calcium, 480 mg/100 g of sodium, 368 mg/100 g of chloride, each.

• Archives of Pharmacal Research
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• 제29권3호
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• pp.241-248
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• 2006

Angiotensin converting enzyme (ACE) inhibitors have cardioprotective effects in different species including human. This cardioprotective effect is mainly due to the inhibition of bradykinin (BK) degradation rather than inhibition of the conversion of angiotensin I to angiotensir. II. Bradykinin, a nonapeptide, has been considered to be the potential target for various enzymes including ACE, neutral endopeptidase 24.11, carboxypeptidase M, carboxypeptidase N, proline aminopeptidase, endopeptidase 24.15, and meprin. In the present study, the coronary vascular beds of Sprague Dawley rat isolated hearts were perfused (single passage) with Krebs solution alone or with different concentrations of BK i.e. $2.75{\times}10^{-10},\;10^{-7},\;10^{-6}\;and\;10^{-5}M$ solution. Percent degradation of BK was determined by radioimmunoassay. The degradation products of BK after passing through the isolated rat-hearts were determined using RP-HPLC and mass spectroscopy. All the four doses of BK significantly decreased the perfusion pressure during their passage through the hearts. The percentage degradation of all four doses was decreased as the concentration of drug was increased, implying saturation of a fixed number of active sites involved in BK degradation. Bradykinin during a single passage through the hearts degraded to give [1-7]-BK as the major metabolite, and [1-8]-BK as a minor metabolite, detected on HPLC. Mass spectroscopy not only confirmed the presence of these two metabolites but also detected traces of [1-5]-BK and arginine. These findings showed that primarily ACE is the major cardiac enzyme involved in the degradation of bradykinin during a single passage through the coronary vascular of bed the healthy rat heart, while carboxypeptidase M may have a minor role.

• 한국수산과학회지
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• 제47권2호
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• pp.135-143
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• 2014

Exopeptidase active fractions from the hepatopancreas of the Argentina shortfin squid Illex argentinus, were obtained with acetone (AC 30-40%), ammonium sulfate (AS 60-70% saturation), anion exchange chromatography (AE-II, 0.2 M NaCl) and gel filtration chromatography (GF-I, 30-50 kDa) fractionation methods. A bitter peptide solution that has a bitterness equivalent to that of 2% glycylphenylalanine and prepared by tryptic hydrolysis of milk casein, was treated with the exopeptidase active fractions. The GF-I fraction was the best based on aminopeptidase activity (35.3 U/mg), percentage of recovery (30.7%) and a sensory evaluation (1.7). The amount of released amino acids increased as incubation time increased, and the bitterness of the enzyme reaction mixtures decreased. Incubation with the GF-I fraction for 24 h resulted in the hydrolysis of several peptides as revealed by the reverse-phase high performance liguid chromatography profile, with three peaks (3, 5 and 6) decreasing in area (%) and three peaks (1, 2 and 4) increasing in area (%). Therefore, the GF-I fraction appeared to be ideally suited to reduce bitterness in protein hydrolysates by catalyzing the hydrolysis of bitter peptides.