• Title/Summary/Keyword: trypsin inhibitor activity

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A Prolyl Endopeptidase-lnhibiting Antioxidant from Phyllanthus ussurensis

  • Chung, Shin-kyo;Nam, Ji-Ae;Jeon, So-Young;Kim, Sang-ln;Lee, Hee-Ju;Chung, Tai-Ho;Song, Kyung-Sik
    • Archives of Pharmacal Research
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    • v.26 no.12
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    • pp.1024-1028
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    • 2003
  • A prolyl endopeptidase inhibitor was isolated from the ethyl acetate soluble fraction of Phyllanthus ussurensis. The active compound was identified as an ellagitannin, corilagin. It was shown to non-competitively inhibit prolyl endopeptidase (PEP) with the $IC_{50}$ value of $1.17 \times $10^{-6}\mu$M. The Ki value was $6.70 \times 10^{-7}$ M. Corilagin was less inhibitory to other serine proteases such as chymotrypsin, trypsin, and elastase, indicating that it was relatively a specific inhibitor of PEP. Corilagin also effectively inhibited reactive oxygen species such as hydroxide and superoxide anion radical, hydrogen peroxide, and DPPH. Especially, corilagin showed potent scavel1ging activity on the superoxide anion radical in the ESR method ($IC_{50} =3.79 \times 10^{-6}$M) as well as xanthine oxidase system.

Purification of Angiotensin I-Converting Enzyme Inhibitory Peptide from Squid Todarodes pacificus Skin (오징어(Todarodes pacificus) 껍질로부터 Angiotensin I 전환효소 저해 펩티드의 분리 정제)

  • Lee, Jung-Kwon;Jeon, Joong-Kyun;Byun, Hee-Guk
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.44 no.2
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    • pp.118-125
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    • 2011
  • In this study, an angiotensin I-converting enzyme (ACE) inhibitor from squid skin was purified and characterized. Squid (Todarodes pacificus) skin protein isolates were hydrolyzed using six commercial proteases: alcalase, ${\alpha}$-chymotrypsin, neutrase, papain, pepsin, and trypsin. The peptic hydrolysate had the highest ACE inhibitory activity. The ACE inhibitory peptide was purified using Sephadex G-25 column chromatography and reverse phase high-performance liquid chromatography (HPLC) with a $C_{18}$ column. The purified ACE inhibitory peptide was identified and sequenced, and found to consist of seven amino acid residues: Ser-Ala-Gly-Ser-Leu-Val-Pro (657Da). The $IC_{50}$ value of the purified ACE inhibitory peptide was 766.2 ${\mu}M$, and Lineweaver-Burk plots suggested that the purified peptide acts as a noncompetitive ACE inhibitor. These results suggest that the ACE inhibitory peptide purified from the peptic hydrolysate of squid skin may be of benefit in developing antihypertensive drugs and functional foods.

Comparative Biochemical Properties of Proteinases from the Hepatopancreas of Shrimp. -I. Purification of Protease from the Hepatopancreas of Penaeus japonicus-

  • Choi Sung-Mi;Oh Eun-Sil;Kim Doo-Sang;Pyeun Jae-Hyeung;Cho Deuk-Moon;Ahn Chang-Bum;Kim Hyeung-Rak
    • 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.

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Fifty C-terminal amino acid residues are necessary for the chaperone activity of DFF45 but not for the inhibition of DFF40

  • Park, Hyun-Ho
    • BMB Reports
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    • v.42 no.11
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    • pp.713-718
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    • 2009
  • Apoptotic DNA fragmentation, the hallmark of apoptosis, is mediated primarily by caspase-activated DFF40 (CAD) nuclease. DFF40 exists as a heterodimer with DFF45 (ICAD), which is a specific chaperone and inhibitor of DFF40 under normal conditions. To understand the mechanism through which the DFF40/DFF45 system is regulated, we analyzed the structural and biochemical properties of apoptotic DNA fragmentation mediated by DFF40/DFF45. Using limited proteolysis, we show that residues 1-281 of DFF45 form a rigid, crystallized domain, whereas the loop formed by residues 277-281 is accessible by trypsin. These results show that the C-terminal helix formed by residues 281-300 is dynamic and necessary for the chaperone activity of DFF45, but not for inhibition of DFF40.

이담자균 효모 Rhodosporidium toruloides에서 Rhodotorucine A에 의한 막단백질 인산화의 저해와 Trigger Peptidase의 관련성

  • 정영기;이태호;류병호
    • Microbiology and Biotechnology Letters
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    • v.24 no.6
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    • pp.641-646
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    • 1996
  • [$\gamma$-$^{32}$P]ATP was used to test phosphorylation of membrane proteins of mating type a cells of heterobasidiomycetous yeast Rhodosporidium toruloides separated by non-denaturing electrophoresis. The phosphoprotein was observed in the membrane proteins. The phosphorylation was inhibited by the pheromone rhodotorucine A (Rh. A) secreted by mating type A of the yeast. Rh. A didn't inhibit the phosphorylation in the presence of a trigger peptidase (TPase) inhibitor, antipain. Partially digested Rh. A by trypsin maintained the phosphorylation inhibitory activity. These results show that TPase activity plays an important role in the transduction of pheromone signal in the yeast.

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Pasteurization Efficiency and Physico-chemical Changes of Soymilk HTST Pasteurized Using Microwaves (두유의 마이크로파 고온단시간 살균시 살균효과 및 이화학적 성분 변화)

  • Kim, Suk-Shin;Lee, Joo-Hee
    • Korean Journal of Food Science and Technology
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    • v.31 no.5
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    • pp.1196-1202
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    • 1999
  • This work was to determine the microbial and physico-chemical changes of HTST-pasteurized soymilk using microwave energy. Soymilk was HTST pasteurized$(at\;90^{\circ}C\;for\;20\;sec)$ by three methods: by heating in a stainless steel tube immersed in a hot water bath(MP0), by heating in a microwave cavity to a defiled temperature and then holding in a hot water bath(MP1), and by both heating and holding in a microwave cavity(MP2). The microbial quality based on the total plate count was in the order of MP0, MP2 and MP1. The three samples pasteurized by different methods showed the similar microbial quality with respect to the coliform count, psychrotrophic bacterial count and phosphatase activity. The destruction of trypsin inhibitor was in the order of MP0, MP1 and MP2. There were no significant differences in pH, titratable acidity, viscosity and vitamin $B_2$ content before and after pasteurization and among the different pasteurization methods. The similar or higher quality retention of the MP1 or MP2 supports the possibility of using microwave energy for the HTST pasteurization of soymilk and other fluid food products.

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Health Functional Peptides from Milk Products (유제품의 기능성 펩타이드)

  • Lee, Hyong-Joo
    • Journal of Dairy Science and Biotechnology
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    • v.16 no.2
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    • pp.98-105
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    • 1998
  • Various peptides derived from food are among the most potent physiologically active agents known, and include anticancer peptides, angiotensin converting enzyme(ACE) inhibitor exhibiting antihypertension action, opioid peptides, antithrombotic peptides, hypocholesterolemic peptides, immunomodulators, calcium absorption enhancers, and other peptides. Hydrophobic peptides extracted from a Cheddar-type cheese slurry were fractionated by gel chromatography and repeated HPLC. A peptide fraction from HPLC showed high cytotoxicity on the tumor cell lines such as a human colon carcinoma, and comprised of Tyr, Ser, Leu, Gly, and others. Hypocholesterolemic peptides were isolated from peptic hydrolyzates of casein and soy proteins. Macropeptides of 1,000${\sim}$5,000 dalton were effective on reducing the cholesterol level of mouse serum. Peptides showing high Krigbaum hydrophobicity and ANS surface hydrophobicity resulted in high hypocholesterolemic effect and fecal steroid concentrations. Caseinomacropeptides(CMP) were isolated from whey powder and treated with soluble and immobilized trypsin to obtain antithrombotic peptides. One fraction from the CMP hydrolyzed with immobilized trypsin for 24h exhibited high antithrombotic activity with 52.5% inhibition of platelet aggregation. These result suggested that peptides from various milk products could be utilized as a good bioactive agents for developing health functional foods.

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Health Functional Peptides From Milk Products (유제품의 기능성 펩타이드)

  • Lee, Hyong-Joo
    • 한국유가공학회:학술대회논문집
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    • 1998.05a
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    • pp.22-29
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    • 1998
  • Various peptides derived from food are among the most potent physiologically active agents known, and include anticancer peptides, angiotensin converting enzyme(ACE) inhibitor exhibiting antihypertension action, opioid peptides, antithrombotic peptides, hypocholesterolemic peptides, immunomodulators, calcium absorption enhancers, and other peptides. Hydrophobic peptides extracted from a Cheddar-type cheese slurry were fractionated by gel chromatography and repeated HPLC. A peptide fraction from HPLC showed high cytotoxicity on the tumor cell lines such as a human colon carcinoma, and comprised of Tyr, Ser, Leu, Gly, and others. Hypocholesterolemic peptides were isolated from peptic hydrolyzates of casein and soy proteins. Macropeptides of 1,000${\sim}$5,000 dalton were effective on reducing the cholesterol level of mouse serum. Peptides showing high Krigbaum hydrophobicity and ANS surface hydrophobicity resulted in high hypocholesterolemic effect and fecal steroid concentrations. Caseinomacropeptides (CMP) were isolated from whey powder and treated with soluble and immobilized trypsin to obtain antithrombotic peptides. One fraction from the CMP hydrolyzed with immobilized trypsin for 24h exhibited high antithrombotic activity with 52.5% inhibition of platelet aggregation. These results suggested that peptides from various milk products could be utilized as a good bioactive agents for developing health functional foods.

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EFFECT OF HEAT TREATMENT ON NUTRITIONAL VALUE OF WINGED BEAN (Psophocarpus tetragonolobus) AS COMPARED TO SOYBEAN I. CHEMICAL CHARACTERISTICS OF TREATED WINGED BEAN

  • Mutia, R.;Uchida, S.
    • Asian-Australasian Journal of Animal Sciences
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    • v.6 no.1
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    • pp.19-26
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    • 1993
  • The effect heat treatment (autoclave) on nutritional value of winged bean as compared to soybean has been investigated. The winged bean and soybean were obtained from local cultivar grown in Indonesia. The beans were autoclaved at $120^{\circ}C$ for 15, 30, 45, 60 or 90 minutes, respectively before being ground for chemical analysis. Trypsin inhibitors of winged bean and soybean decreased (p < 0.05) along with decreasing of urease activity as heating time increased from 0 to 90 minutes. Heat treatment significantly (p < 0.05) reduced protein solubility in 0.2% potassium hydroxide of winged bean as well as soybean. In vitro protein digestibility was significantly (p < 0.05) improved by heating treatment (15 to 60 min of autoclaving), however, excessive heating (90 min of autoclaving) decreased the digestibility of winged beans. Excessive heating had adverse effect on lysine, cystine and methionine contents of winged beans. The results of this study suggested that autoclaving at $120^{\circ}C$ within 45 minutes should be adequate to remove protease inhibitors and could improve protein digestibility of winged beans.

Screening System for Chitin Synthase II Inhibitors from Natural Resources and its Inhibitor Prodigiosin

  • Hwang, Eui-Il;Kim, Young-Kook;Lee, Hyang-Bok;Kim, Hong-Gi;Kim, Sung-Uk
    • Journal of Microbiology and Biotechnology
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    • v.10 no.2
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    • pp.251-257
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    • 2000
  • Chitin synthases are identified as key enzymes of chitin biosynthesis in most of the fungi. Among them, chitin synthase II has been reported to be and essential enzyme in chitin biosynthesis, and exists as a membrane-bound form. To search and screen new antifungal agents from natural resources to inhibit chitin synthase II, the assay conditions were established using the enzyme isolated from Saccharomyces cerevisiae ECY38-38A(pAS6) that overproduces only chitin synthase II. This enzyme was activated only by partial proteolysis with trypsin. Its actibity reached the maximum at $80{\;}\mu\textrm{g}/ml$ of trypsin and was strongly stimulated by 2.0 mM $Co^{2+}$, 1.0 nM UDP-[$^{14}C$]-GicNAc, and 32 mM free-GlcNAc. Under these assay conditions, the highest chitin synthase II activity was observed by incubation at $30^{\circ}C$ for 90 min. However, and extremely narrow range of organic solvents up to as much as 25% of DMSO and 25% of MeOH was useful for determining optimal assay conditions. After a search or potent inhibitors of chitin synthase II from natural resources, prodigiosin was isolated from Serratia marcescens and purified by solvent extration and silica gel column chromatographies. The structure of prodigiosin was determined by UV, IR, Mass spectral, and NMR spectral analyses. Its molecular weight and formula were found to be 323 and $C_{20}H_{25}N_{3}O$, respectively. Prodigiosin ingibited chitin synthase II by 50% at the concentration of $115{\;}\mu\textrm{g}/ml$.

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