[Objective] The aim of this study was to study effects of metal ions on the protease activities in digestive tissues and gland of red-white ornamental carp（Cyprinus carpio L）.[Method] Effects of four kinds of metal ...[Objective] The aim of this study was to study effects of metal ions on the protease activities in digestive tissues and gland of red-white ornamental carp（Cyprinus carpio L）.[Method] Effects of four kinds of metal ions （K＋,Na＋,Mg2＋ and Ca2＋） on protease activities in hepatopancreas,foregut,midgut,hindgut of red-white ornamental carp were studied by enzyme analysis method.[Result] Effects of four kinds of metal ions on protease activities of red-white ornamental carp were different in the range of experimental concentration from 25 mmol/L to 150 mmol/L.K＋ could promote protease activities in hepatopancreas and hindgut at different levels.Especially,K＋ had the promoting effect at low-concentration level,but the inhibitory effect at high-concentration level in midgut and the inhibitory effect in foregut.Na＋ had the promoting effect on protease activities in hepatopancreas,foregut and hindgut at different levels,but the inhibitory effect in midgut.Mg2＋ and Ca2＋ had the inhibitory effect on protease activities in intestinal and hepatopancreas at different levels.[Conclusion] This study provides basic data and theoretical foundation for researches on the digestive physiology of red-white ornamental carp or the development and optimization of compound feed.展开更多
Previously, we have purified Jerdonitin from Trimeresurusjerdonii venom. Compared with other P-Ⅱ class snake venom metalloproteinases （SVMPs）, Jerdonitin has a primary structure comprising metalloproteinase and dis...Previously, we have purified Jerdonitin from Trimeresurusjerdonii venom. Compared with other P-Ⅱ class snake venom metalloproteinases （SVMPs）, Jerdonitin has a primary structure comprising metalloproteinase and disintegrin domains. However, no hemorrhagic and fibrinogenolytic activities were detected for Jerdonitin. We thought that organic buffer of high performance liquid charamatography （HPLC） might affect its enzymatic activity. In this study, we purified Jerdonitin by another procedure excluding the HPLC. It was homogenous as judged by SDS-PAGE and had an apparent molecular weight of 36 kDa under non-reducing conditions and 38 kDa under reducing conditions, respectively. Like other typical metalloproteinases, Jerdonitin preferentially degraded alpha-chain of human fibrinogen and this fibrinogenolytic activity was completely inhibited by EDTA, but not by PMSF. It was interesting that Jerdonitin did not induce hemorrhage after intradermal injection in mice.展开更多
A neutral metalloprotease was purified from the cultured mycelia of Laccocephalum mylittae,an effective medicinal fungus widely used in anthelmintic therapy.The protease was purified to homogeneity with 31.85-fold pur...A neutral metalloprotease was purified from the cultured mycelia of Laccocephalum mylittae,an effective medicinal fungus widely used in anthelmintic therapy.The protease was purified to homogeneity with 31.85-fold purification and a final yield of 21.76%.The subunit molecular weight of the protease is about 40000 estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE).The optimum reaction pH and temperature are 7.5 and 50oC,respectively.The protease activity is largely enhanced by Ca2 +,but highly inhibited by tetrasodium ethylenediaminetetraacetate(EDTA),a metal-chelator,suggesting that the enzyme is a metalloprotease.The Michaelis-Menten constan Km and Vmax value for casein substrate are 6.09 mg·ml -1and 21.32μg·min -1·ml -1, respectively.In vitro anthelmintic tests of the protease exhibit distinct lethal effects on the third stage larvae(L3)of Ascaris suum.Scanning electron microscopy and SDS-PAGE analysis indicates that the proteolysis of larvae proteins caused by this protease may relate to the anthelmintic activity of L.mylittae.展开更多
We cloned and sequenced a prtV-like gene from Vibrio anguillarum M3 strain.This prtV gene encodes a putative protein of 918 amino acids,and is highly homologous to the V.cholerae prtV gene.We found that a prtV inserti...We cloned and sequenced a prtV-like gene from Vibrio anguillarum M3 strain.This prtV gene encodes a putative protein of 918 amino acids,and is highly homologous to the V.cholerae prtV gene.We found that a prtV insertion mutant strain displayed lower gelatinase activity on gelatin agar,lower protease activity against azocasein,and lower activity for four glycosidases.This prtV mutant strain also had increased activity for two esterases in its extracellular products,as analyzed by the API ZYM system.In addition,the prtV mutant strain exhibited decreased growth in turbot intestinal mucus and reduced hemolytic activity on turbot erythrocytes.Infection experiments showed that the LD50 of the prtV mutant strain increased by at least 1 log compared to the wild-type in turbot fish.We propose that prtV plays an important role in the pathogenesis of V.anguillarum.展开更多
Effects of genistein on invasion and matrix metalloproteinase activities were investigated in HT1080 human sarcoma cells.Invasion of HT1080 cells through reconstituted basement membrane was inh...Effects of genistein on invasion and matrix metalloproteinase activities were investigated in HT1080 human sarcoma cells.Invasion of HT1080 cells through reconstituted basement membrane was inhibited when the cells were treated with 100 μ mol/L and 200 μ mol/L genistein.At the same concentrations,genistein not only suppressed latent forms of matrix metalloprotinese 2 and 9(MMP 2 and MMP 9) to convert into active forms,but also increase dramatically the tissue inhibitor of metalloproteinase(TIMP 1) mRNA contents and reverse the imbalance of MMPs and TIMPs.However,expressions of MMP 2 and MMP 9 were not significantly affected.Suppression of MMP activation and increase of TIMP 1 expression will decrease matrix degradation by MMPs,and consequently inhibit invasions of the cells.These results emphasized the existence of the imbalance between MMPs and TIMPs in tumor invasion and metastasis formation.The value of genistein as a drug for antiinvasion and anti metastasis chemotherapy was suggested.展开更多
文摘[Objective] The aim of this study was to study effects of metal ions on the protease activities in digestive tissues and gland of red-white ornamental carp（Cyprinus carpio L）.[Method] Effects of four kinds of metal ions （K＋,Na＋,Mg2＋ and Ca2＋） on protease activities in hepatopancreas,foregut,midgut,hindgut of red-white ornamental carp were studied by enzyme analysis method.[Result] Effects of four kinds of metal ions on protease activities of red-white ornamental carp were different in the range of experimental concentration from 25 mmol/L to 150 mmol/L.K＋ could promote protease activities in hepatopancreas and hindgut at different levels.Especially,K＋ had the promoting effect at low-concentration level,but the inhibitory effect at high-concentration level in midgut and the inhibitory effect in foregut.Na＋ had the promoting effect on protease activities in hepatopancreas,foregut and hindgut at different levels,but the inhibitory effect in midgut.Mg2＋ and Ca2＋ had the inhibitory effect on protease activities in intestinal and hepatopancreas at different levels.[Conclusion] This study provides basic data and theoretical foundation for researches on the digestive physiology of red-white ornamental carp or the development and optimization of compound feed.
文摘Previously, we have purified Jerdonitin from Trimeresurusjerdonii venom. Compared with other P-Ⅱ class snake venom metalloproteinases （SVMPs）, Jerdonitin has a primary structure comprising metalloproteinase and disintegrin domains. However, no hemorrhagic and fibrinogenolytic activities were detected for Jerdonitin. We thought that organic buffer of high performance liquid charamatography （HPLC） might affect its enzymatic activity. In this study, we purified Jerdonitin by another procedure excluding the HPLC. It was homogenous as judged by SDS-PAGE and had an apparent molecular weight of 36 kDa under non-reducing conditions and 38 kDa under reducing conditions, respectively. Like other typical metalloproteinases, Jerdonitin preferentially degraded alpha-chain of human fibrinogen and this fibrinogenolytic activity was completely inhibited by EDTA, but not by PMSF. It was interesting that Jerdonitin did not induce hemorrhage after intradermal injection in mice.
基金Supported by the National High Technology Research and Development Program of China（2007AA021506） the Natural Science Foundation of Zhejiang Province（R207609）
文摘A neutral metalloprotease was purified from the cultured mycelia of Laccocephalum mylittae,an effective medicinal fungus widely used in anthelmintic therapy.The protease was purified to homogeneity with 31.85-fold purification and a final yield of 21.76%.The subunit molecular weight of the protease is about 40000 estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE).The optimum reaction pH and temperature are 7.5 and 50oC,respectively.The protease activity is largely enhanced by Ca2 +,but highly inhibited by tetrasodium ethylenediaminetetraacetate(EDTA),a metal-chelator,suggesting that the enzyme is a metalloprotease.The Michaelis-Menten constan Km and Vmax value for casein substrate are 6.09 mg·ml -1and 21.32μg·min -1·ml -1, respectively.In vitro anthelmintic tests of the protease exhibit distinct lethal effects on the third stage larvae(L3)of Ascaris suum.Scanning electron microscopy and SDS-PAGE analysis indicates that the proteolysis of larvae proteins caused by this protease may relate to the anthelmintic activity of L.mylittae.
基金Supported by the National Basic Research Program of China (973 Program) (No. 2006CB101803)the National Natural Science Foundation of China (No. 30871935)the National High Technology Research and Development Program (863 Program) (No. 2003AA622070)
文摘We cloned and sequenced a prtV-like gene from Vibrio anguillarum M3 strain.This prtV gene encodes a putative protein of 918 amino acids,and is highly homologous to the V.cholerae prtV gene.We found that a prtV insertion mutant strain displayed lower gelatinase activity on gelatin agar,lower protease activity against azocasein,and lower activity for four glycosidases.This prtV mutant strain also had increased activity for two esterases in its extracellular products,as analyzed by the API ZYM system.In addition,the prtV mutant strain exhibited decreased growth in turbot intestinal mucus and reduced hemolytic activity on turbot erythrocytes.Infection experiments showed that the LD50 of the prtV mutant strain increased by at least 1 log compared to the wild-type in turbot fish.We propose that prtV plays an important role in the pathogenesis of V.anguillarum.
文摘Effects of genistein on invasion and matrix metalloproteinase activities were investigated in HT1080 human sarcoma cells.Invasion of HT1080 cells through reconstituted basement membrane was inhibited when the cells were treated with 100 μ mol/L and 200 μ mol/L genistein.At the same concentrations,genistein not only suppressed latent forms of matrix metalloprotinese 2 and 9(MMP 2 and MMP 9) to convert into active forms,but also increase dramatically the tissue inhibitor of metalloproteinase(TIMP 1) mRNA contents and reverse the imbalance of MMPs and TIMPs.However,expressions of MMP 2 and MMP 9 were not significantly affected.Suppression of MMP activation and increase of TIMP 1 expression will decrease matrix degradation by MMPs,and consequently inhibit invasions of the cells.These results emphasized the existence of the imbalance between MMPs and TIMPs in tumor invasion and metastasis formation.The value of genistein as a drug for antiinvasion and anti metastasis chemotherapy was suggested.
