The chemical synthesis of Guanine arabinoside （ara-G） is extremely complex, time-consuming, and seriously polluted. A two-step enzymatic synthesis process was developed to acquire ara-G easily. 2,6-Diaminopurine ara...The chemical synthesis of Guanine arabinoside （ara-G） is extremely complex, time-consuming, and seriously polluted. A two-step enzymatic synthesis process was developed to acquire ara-G easily. 2,6-Diaminopurine arabinoside （ara-DA） was first synthesized with purine nucleoside phosphorylase and pyrimidine nucleoside phosphorylase produced by Enterobacter aerogenes DGW-07. The conversion yield of ara-DA could reach above 90% when the reaction liquid contained 30 mmol·L^-1 uracil arabinoside as arabinose donor, 10 mmol·L^- 1 2,6-diaminopurine as arabinose acceptor in pH 7.0 20 mmol·L^-1 phosphate buffer, and reacted at 60℃ for 48h. Then, ara-DA was effectively transformed into ara-G with adenylate deaminase produced by Aspergillus oryzae DAW-01. The total process had no complex separation and purification.展开更多
Alkaline phosphatases(APs) are non-specifi c phosphohydrolases, and they are widely used in clinical diagnostics and biological studies. APs are widespread in nature and exhibit dif ferent structural formulations. Bas...Alkaline phosphatases(APs) are non-specifi c phosphohydrolases, and they are widely used in clinical diagnostics and biological studies. APs are widespread in nature and exhibit dif ferent structural formulations. Based on the diversity of biogenetic sources, APs exhibit temperature-propensity traits, and they are classifi ed as psychrophilic, mesophilic, and thermophilic. In this article, the characteristics of psychrophilic APs from marine organisms were described, accompanied by a simple description of APs from other organisms. This review will facilitate better utilization of marine APs in the biotechnology fi eld.展开更多
AIM:To investigate the effect of chloride intracellular channel 1(CLIC1) on the cell proliferation,apoptosis,migration and invasion of gastric cancer cells.METHODS:CLIC1 expression was evaluated in human gastric cance...AIM:To investigate the effect of chloride intracellular channel 1(CLIC1) on the cell proliferation,apoptosis,migration and invasion of gastric cancer cells.METHODS:CLIC1 expression was evaluated in human gastric cancer cell lines SGC-7901 and MGC-803 by real time polymerase chain reaction(RT-PCR).Four segments of small interference RNA(siRNA) targeting CLIC1 mRNA and a no-sense control segment were designed by bioinformatics technology.CLIC1 siRNA was selected using Lipofectamine 2000 and transfected transiently into human gastric cancer SGC-7901 and MGC-803 cells.The transfected efficiency was observed under fluorescence microscope.After transfection,mRNA expression of CLIC1 was detected with RT-PCR and Western blotting was used to detect the protein expression.Proliferation was examined by methyl thiazolyl tetrazolium and apoptosis was detected with flow cytometry.Polycarbonate membrane transwell chamber and Matrigel were used for the detection of the changes of invasion and migration of the two cell lines.RESULTS:In gastric cancer cell lines SGC-7901 and MGC-803,CLIC1 was obviously expressed and CLIC1 siRNA could effectively suppress the expression of CLIC1 protein and mRNA.Proliferation of cells transfected with CLIC1 siRNA3 was enhanced notably,and the highest proliferation rate was 23.3%(P = 0.002) in SGC-7901 and 35.55%(P = 0.001) in MGC-803 cells at 48 h.The G2/M phase proportion increased,while G0/G1 and S phase proportions decreased.The apoptotic rate of the CLIC1 siRNA3 group obviously decreased in both SGC-7901 cells(62.24%,P = 0.000) and MGC-803 cells(52.67%,P = 0.004).Down-regulation of CLIC1 led to the inhibition of invasion and migration by 54.31%(P = 0.000) and 33.62%(P = 0.001) in SGC-7901 and 40.74%(P = 0.000) and 29.26%(P = 0.002) in MGC-803.However,there was no significant difference between the mock group cells and the negative control group cells.展开更多
An antithrombus enzyme (ATE) was precipitated by (NH4)2SO4 or ethanol from supernatant of Bacillus subtilis culture broth then purified using ion exchange chromatography on CM-sepharose fast flow. The effects of ionic...An antithrombus enzyme (ATE) was precipitated by (NH4)2SO4 or ethanol from supernatant of Bacillus subtilis culture broth then purified using ion exchange chromatography on CM-sepharose fast flow. The effects of ionic strength and pH value on protein adsorption, the gradient elution at different flow rates and step elution were examined respectively. The recovery yield of the optimised process was 74.5% with a purification factor 8.1. The ATE molecular weight was estimated as 30ku by SDS-PAGE. The experimental results showed that the enzyme was stable in the range of pH 7 to pH11, and temperature 25℃ to 37℃.展开更多
Plasma membrane of plant cells is surrounded by cellulose wall and adjacent cells are joined together by a thick pectin rich matrix. Separation of plant cells and removal of the cell wall experimentally, by either a m...Plasma membrane of plant cells is surrounded by cellulose wall and adjacent cells are joined together by a thick pectin rich matrix. Separation of plant cells and removal of the cell wall experimentally, by either a mechanical or an enzymatic process, results in the production ofprotoplast. Protoplasts are useful tools to study the uptake and transport ofmacromolecules and production of somatic hybrids. Protoplasts can be obtained from all types of actively growing young and healthy tissues. The most convenient and widely used source of plant protoplasts is leaf. Juvenile seedling tissues, cotyledons are other alternative tissues most frequently used for protoplasts isolation. All the environmental and genotypic factors, which affect the cell wall thickenings and compactness indirectly, influence the number of protoplasts recovered. Protoplasts are isolated by two methods, mechanical and enzymatic. The enzyme mixture solution of celluiose/macerozyme is used to digest the cell wall. The critical factors affecting the obtaning ofprotoplasts are the kinds of cell wall degrading enzymes, the physiological state of plant leaves, the type of osmotic stabilizers and the composition of reaction solution. With the improvement of technique and enzyme combination rate, the yield of collected protoplasts will be increased higher.展开更多
With the development of colloid interface and enzyme technologies,enzyme-containing reversed micellar system has been receiving much attention in bioseparation and bioconversion. Because of its high efficiency,it has ...With the development of colloid interface and enzyme technologies,enzyme-containing reversed micellar system has been receiving much attention in bioseparation and bioconversion. Because of its high efficiency,it has brought new opportunities for the development of molecular biotechnology. Reversed micelles represent nano-sized aqueous droplets stabilized by surfactant amphiphiles inside the bulk organic solvents. The entrapped enzymes have enhanced activities under those conditions as suited in the lipid bilayers of biological membranes. The fundamentals of enzyme-containing reversed micellar system are described in this paper,with special emphasis on the effects of surfactants varying in concentrations and structures. The latest study progress on the surfactants application in enzyme-containing reversed micelles is reviewed. The introduction of novel functional surfactants in micellar enzymology and their future development are also discussed.展开更多
基金Supported by the Innovation Fund for Technology Based Firms from Ministry of Science and Technology of China(07C26213101283)
文摘The chemical synthesis of Guanine arabinoside （ara-G） is extremely complex, time-consuming, and seriously polluted. A two-step enzymatic synthesis process was developed to acquire ara-G easily. 2,6-Diaminopurine arabinoside （ara-DA） was first synthesized with purine nucleoside phosphorylase and pyrimidine nucleoside phosphorylase produced by Enterobacter aerogenes DGW-07. The conversion yield of ara-DA could reach above 90% when the reaction liquid contained 30 mmol·L^-1 uracil arabinoside as arabinose donor, 10 mmol·L^- 1 2,6-diaminopurine as arabinose acceptor in pH 7.0 20 mmol·L^-1 phosphate buffer, and reacted at 60℃ for 48h. Then, ara-DA was effectively transformed into ara-G with adenylate deaminase produced by Aspergillus oryzae DAW-01. The total process had no complex separation and purification.
基金Supported by the National Natural Science Foundation for Creative Research Groups(No.41221004)the Natural Science Foundation of Shandong Province(No.ZR2011DQ005)+6 种基金the Key Laboratory of Marine Ecology and Environmental Science and Engineering,SOA(No.MESE-2013-03)the Key Laboratory for Ecological Environment in Coastal Areas,State Oceanic Administration(No.201306)the Major International Joint Research Project of NSFC(No.41320104008)the Changjiang Scholars Program,Ministry of Education of Chinathe Taishan Scholars Program of Shandong Provincethe Key Lab of Marine Bioactive Substance and Modern Analytical Technique,SOA(No.MBSMAT-2013-05)This is MCTL Contribution No.81
文摘Alkaline phosphatases(APs) are non-specifi c phosphohydrolases, and they are widely used in clinical diagnostics and biological studies. APs are widespread in nature and exhibit dif ferent structural formulations. Based on the diversity of biogenetic sources, APs exhibit temperature-propensity traits, and they are classifi ed as psychrophilic, mesophilic, and thermophilic. In this article, the characteristics of psychrophilic APs from marine organisms were described, accompanied by a simple description of APs from other organisms. This review will facilitate better utilization of marine APs in the biotechnology fi eld.
基金Supported by The National Natural Science Foundation of China,No.30560151the Key Research Project of Guangxi Municipal Health Bureau,No.200824+1 种基金the Research Project of Guangxi Educational Department,No.201012MS062 and No. 2011105981002M204the Natural Science Foundation of Guangxi,No.0832113
文摘AIM:To investigate the effect of chloride intracellular channel 1(CLIC1) on the cell proliferation,apoptosis,migration and invasion of gastric cancer cells.METHODS:CLIC1 expression was evaluated in human gastric cancer cell lines SGC-7901 and MGC-803 by real time polymerase chain reaction(RT-PCR).Four segments of small interference RNA(siRNA) targeting CLIC1 mRNA and a no-sense control segment were designed by bioinformatics technology.CLIC1 siRNA was selected using Lipofectamine 2000 and transfected transiently into human gastric cancer SGC-7901 and MGC-803 cells.The transfected efficiency was observed under fluorescence microscope.After transfection,mRNA expression of CLIC1 was detected with RT-PCR and Western blotting was used to detect the protein expression.Proliferation was examined by methyl thiazolyl tetrazolium and apoptosis was detected with flow cytometry.Polycarbonate membrane transwell chamber and Matrigel were used for the detection of the changes of invasion and migration of the two cell lines.RESULTS:In gastric cancer cell lines SGC-7901 and MGC-803,CLIC1 was obviously expressed and CLIC1 siRNA could effectively suppress the expression of CLIC1 protein and mRNA.Proliferation of cells transfected with CLIC1 siRNA3 was enhanced notably,and the highest proliferation rate was 23.3%(P = 0.002) in SGC-7901 and 35.55%(P = 0.001) in MGC-803 cells at 48 h.The G2/M phase proportion increased,while G0/G1 and S phase proportions decreased.The apoptotic rate of the CLIC1 siRNA3 group obviously decreased in both SGC-7901 cells(62.24%,P = 0.000) and MGC-803 cells(52.67%,P = 0.004).Down-regulation of CLIC1 led to the inhibition of invasion and migration by 54.31%(P = 0.000) and 33.62%(P = 0.001) in SGC-7901 and 40.74%(P = 0.000) and 29.26%(P = 0.002) in MGC-803.However,there was no significant difference between the mock group cells and the negative control group cells.
文摘An antithrombus enzyme (ATE) was precipitated by (NH4)2SO4 or ethanol from supernatant of Bacillus subtilis culture broth then purified using ion exchange chromatography on CM-sepharose fast flow. The effects of ionic strength and pH value on protein adsorption, the gradient elution at different flow rates and step elution were examined respectively. The recovery yield of the optimised process was 74.5% with a purification factor 8.1. The ATE molecular weight was estimated as 30ku by SDS-PAGE. The experimental results showed that the enzyme was stable in the range of pH 7 to pH11, and temperature 25℃ to 37℃.
文摘Plasma membrane of plant cells is surrounded by cellulose wall and adjacent cells are joined together by a thick pectin rich matrix. Separation of plant cells and removal of the cell wall experimentally, by either a mechanical or an enzymatic process, results in the production ofprotoplast. Protoplasts are useful tools to study the uptake and transport ofmacromolecules and production of somatic hybrids. Protoplasts can be obtained from all types of actively growing young and healthy tissues. The most convenient and widely used source of plant protoplasts is leaf. Juvenile seedling tissues, cotyledons are other alternative tissues most frequently used for protoplasts isolation. All the environmental and genotypic factors, which affect the cell wall thickenings and compactness indirectly, influence the number of protoplasts recovered. Protoplasts are isolated by two methods, mechanical and enzymatic. The enzyme mixture solution of celluiose/macerozyme is used to digest the cell wall. The critical factors affecting the obtaning ofprotoplasts are the kinds of cell wall degrading enzymes, the physiological state of plant leaves, the type of osmotic stabilizers and the composition of reaction solution. With the improvement of technique and enzyme combination rate, the yield of collected protoplasts will be increased higher.
基金supported by the National Natural Science Foundation of China (50978087,50978088 and 51039001)the Program for Changjiang Scholars and Innovative Research Team in University (IRT0719)+1 种基金the Hu-nan Provincial Natural Science Foundation of China (10JJ7005)the Hunan Key Scientific Research Project (2009FJ1010)
文摘With the development of colloid interface and enzyme technologies,enzyme-containing reversed micellar system has been receiving much attention in bioseparation and bioconversion. Because of its high efficiency,it has brought new opportunities for the development of molecular biotechnology. Reversed micelles represent nano-sized aqueous droplets stabilized by surfactant amphiphiles inside the bulk organic solvents. The entrapped enzymes have enhanced activities under those conditions as suited in the lipid bilayers of biological membranes. The fundamentals of enzyme-containing reversed micellar system are described in this paper,with special emphasis on the effects of surfactants varying in concentrations and structures. The latest study progress on the surfactants application in enzyme-containing reversed micelles is reviewed. The introduction of novel functional surfactants in micellar enzymology and their future development are also discussed.
