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Purification, Immobilization and Characterization of Lipase isoenzyme from <i>Aspergillus niger</i>with C8 Magnetic Particles

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Advances in Bioscience and Biotechnology 2014年第7期5卷 633-641页

作者： Samanta Hernández-García María Inmaculada García-García Francisco García-Carmona Department of Biochemistry and Molecular Biology A Faculty of Biology Regional Campus of International Excellence “Campus Mare Nostrum” University of Murcia Campus Espinardo Murcia Spain

The purification of a lipase isoenzyme from an Aspergillus niger lipase A is reported in this manuscript. Purification was carried out in a simple adsorption step, in which the lipase was offered at low ionic strength to the commercially available C8 modified magnetic particles, MaKProt C8. When the isoenzyme was desorbed with a 0.2% solution of Triton X-100, the SDS-PAGE gel showed a single pure band with a molecular weight of 35 KDa. The purified fraction showed 66.75-fold purification compared with the crude extract. The pure fraction was characterized along with the crude extract and the lipase adsorbed on the MaKProt C8. The purified and the adsorbed lipase showed better activity for the tested substrates (p-nitrophenyl acetate, decanoate, myristate and palmitate) than the crude extract, the preferred substrates being myristate (26.7 μmol·min-1·mg-1) and decanoate (17.4 μmol·min-1·mg-1), respectively. The temperature and pH profiles showed no change for the three enzymes, the optimum temperature being 37°C and the best pH 7.0.

关键词： Protein Purification Protein Affinity Magnetic Hydrophobic Particles Protein Immobilization Lipase Purification Lipase isoenzyme C8 Magnetic Particles

GENETIC DIVERSITY IN THE WILD POPULATION AND HATCHERY STOCK OF Penaeus japonicus SHRIMP BY isoenzyme ANALYSIS

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Zoological Research 2000年第4期21卷 323-326页

作者：庄志猛孟宪红权洁霞戴继勋青岛海洋大学青岛266003 黄海水产研究所青岛266071

Penaeid shrimp Penaeus japonicus,commonly named as Flowery Shrimp or Banded Shrimp,is one of the most commercially important species of marine crustacean fishery in *** of the warm-water species conducting short distance migration,the animal is mainly distributed in the coastal waters of south China,*** southern part of the East China Sea and the South China Sea (Liu et al.,1986).

关键词： Genetic diversity Penaeus japonicus isoenzyme Polyacrylamide gel electrophoresis

Two Human ACAT2 mRNA Variants Produced by Alternative Splicing and Coding for Novel isoenzymes

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Acta Biochimica et Biophysica Sinica 2005年第12期37卷 797-806页

作者： Xiao-Min YAO~(1,2) Can-Hua WANG~(1,2) Bao-Liang SONG~1 Xin-Ying YANG~1 Zhen-Zhen WANG~1 Wei QI~1 Zhi-Xin LIN~2 Catherine C.Y.CHANG~3 Ta-Yuan CHANG~3 Bo-Liang LI~11 State Key Laboratory of Molecular Biology,Institute of Biochemistry and Cell Biology,Shanghai Institutes for Biological Sciences,Chinese Aeademy of Sciences.Shanghai 200031.China:2 Department of Biochemistry and Technology,Shanghai Jiaotong University,Shanghai 200030.China 3 Department of Biochemistry,Dartmouth Medical School.Hanover.New Hampshire 03756.USA State Key Laboratory of Molecular Biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of SciencesShanghai 200031 China Department of Biochemistry and Technology Shanghai Jiaotong UniversityShanghai 200030 China Department of Biochemistry Dartmouth Medical SchoolHanover New Hampshire 03756 USA

Acyl coenzyme A:cholesterol acyltransferase 2 (ACAT2) plays an important role in *** ACAT2 is highly expressed in small intestine and fetal liver,but its expression is greatlydiminished in adult *** full-length human ACAT2 mRNA encodes a protein,designated ACAT2a,with522 amino *** have previously reported the organization of the human ACAT2 gene and the differen-tiation-dependent promoter activity in intestinal Caco-2 *** the current work,two human ACAT2 mRNAvariants produced by alternative splicing are cloned and predicted to encode two novel ACAT2 isoforms,named ACAT2b and ACAT2c,with 502 and 379 amino acids,*** mRNA variants differ fromACAT2a mRNA by lack of the exon 4 (ACAT2b mRNA) and exons 4-5 plus 8-9-10 (ACAT2c mRNA).Significantly,comparable amounts of the alternatively spliced ACAT2 mRNA variants were detected by RT-PCR,and Western blot analysis confirmed the presence of their corresponding proteins in human liver andintestine ***,phosphorylation and enzymatic activity analyses demonstrated that the novelisoenzymes ACAT2b and ACAT2c lacked the phosphorylatable site SLLD,and their enzymatic activitiesreduced to 25%-35% of that of *** evidences indicate that alternative splicing produces twohuman ACAT2 mRNA variants that encode the novel ACAT2 *** findings might help to under-stand the regulation of the ACAT2 gene expression under certain physiological and pathological conditions.

关键词： ACAT2 gene alternative splicing ACAT2 mRNA variant isoenzyme

isoenzymes and soluble protein in Arthrospira from alka-line lakes in Erdos Plateau, China and in exotic species

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Chinese Journal of Oceanology and Limnology 2006年第2期24卷 134-141页

作者：杨茜栗淑媛扈瑞平刘燕乔辰 Common Courses Department Nei Monggo Medical CollegeHohhot 010059 China Department of Biology Nei Monggo Normal UniversityHohhot 010022 China Department of Agronomy Nei Monggo Agricultural University Hohhot 010019 China

The authors compared isoenzymes of five enzymes and soluble protein in Arthrospira platensis (A3) and A. erdosensis (A4) from alkaline lakes in Erdos Plateau, Nei Monggo (Inner Mongalia), China and exotic species of A. platensis (A1) from Chad and A. maxima (A2) from Mexico by means of polyacrylamide gel electrophoresis. The results showed that the isoenzymes of EST, POD and soluble protein were polymorphic. Monomorphism and polymorphism were found in isoenzymes of AMY, CAT and SOD, and monomorphism was found only in the introduced species. The isoenzymes and soluble protein of the local species are all polymorphic. The number of bands in these species were in the order of A3>A4>A1>A2. A2 is the most primary, A1 from Chad Lake is relatively primary, A3 and A4 are ad- vanced species in evolution. Cluster analysis showed that the relation between the two introduced species are the closest to each other, and so too are the two local ones.

关键词： Arthrospira isoenzyme polymorphism cluster analysis

Purification of Active Peroxidase isoenzymes and Their Responses to Nitrogen Fertilization and Rotation of Biomass Sorghum

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American Journal of Plant Sciences 2012年第10期3卷 1422-1429页

作者： Jason P. Wight Frank M. Hons Sanique M. South Godson O. Osuji Biotechnology and Biochemistry Laboratory CARC Prairie View A&M University Prairie View USA Department of Soil and Crop Sciences Texas A&M University College Station USA

Peroxidases (EC 1.***.1.7) participate in lignin biosynthesis. But peroxidation is not a tool for assaying lignocellulose metabolism because the active cannot yet be separated from the inactive peroxidases. A biochemical tool for assaying plant cell wall responses to agronomic practices is needed in the lignocellulosic feedstock renewable energy industry. Peroxidase of biomass sorghum was purified to 9 - 13 charge isomers by free solution IEF (Rotofor) technique. Free solution IEF was more effective than chromatographic purification of active peroxidase isoenzymes. Native PAGE separated each charge isomer to three anionic and three cationic isoenzymes. Hydrogen peroxide and o-dianisidine assays showed that only 20% - 30% of the isoenzymes displayed normal Michaelis-Menten kinetics. Sorghum planted without nitrogen fertilization induced the hydrogen peroxide noncompetitive inhibition of peroxidase, but 280 kg·ha–1 nitrogen fertilization and 100% sorghum mineral residue return to the soil tripled the concentration of active peroxidase and relieved the inhibition with concomitant increases of 350 kg lignin and 3532 kg·cellulose·ha–1. Nitrogen fertilization without crop rotation induced hydrogen peroxide inhibition of the peroxidase, but nitrogen fertilization and 25% sorghum rotation changed the PI of the active peroxidase from neutral to mildly acidic and relieved the inhibition with concomitant enormous increases of 690 kg lignin and 7151 kg·cellulose·ha–1. Hydrogen peroxide inhibition kinetics is consistent with the known peroxidase-substrate intermediate dead-end complex formation. Lignocellulosic yield was greatest under the agronomic management that combined 280 kg·ha–1 nitrogen fertilizer with 25% sorghum residue, which resulted in a shift of pI value of the active peroxidase due to a reduction in the Km value of the peroxidase. Therefore, up to 75% of sorghum biomass rather than only 50% can be harvested for conversion to bioenergy products.

关键词： Oxidoreduction isoenzyme Patterns Inhibition by Hydrogen Peroxide Lignocellulose Yield Sorghum-Sorghum Rotation

A STUDY ON ALKALINE PHOSPHATASE isoenzymeS ASSOCIATED WITH GLUCOCORTICOID HEPATOPATHY IN THE DOG

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A STUDY ON ALKALINE PHOSPHATASE ISOENZYMES ASSOCIATED WITH G...

Effects of UV-B irradiation on isoforms of antioxidant enzymes and their activities in red alga Grateloupia filicina(Rhodophyta)

面向21世纪的动物科学与动物医学国际研讨会

作者： Akira Hashimoto Michelo Syakalima Takiguchi Mitsuyoshi Morita Yutaka Jun Yasuda Department of Veterinary Clinical Sciences Graduate School of Veterinary Medicine Department of Veterinary Clinical Sciences Graduate School of Veterinary Medicine Department of Veterinary Clinical Sciences Graduate School of Veterinary Medicine Department of Veterinary Clinical Sciences Graduate School of Veterinary Medicine Veterinary Teaching Hospital Faculty of Agriculture Iwate University

＜正＞Glucocortlcold hepatopathy（GH） is known in dogs with endogenous hypercortisollsm and in dogs receiving exogenous glucocorticoids. The hepatopathy is characterized histologically by a pattern of cytoplasmic vacuolization and biochemically by induction of liver enzymes, especially the corticosteroid-induced alkaline phosphates（CALP） that is unique to dogs. We investigated the relationship between serum ALP isoenzymes and the degree of GH. The degree of hepatopathy was quantified using vacuolization scores under light microscopy and degree of hepatopathy was quantified using vacuolization scores under light microscopy and digital analysis of hepatic parenchymal histograms using a ultrasonographic electronic scanner

关键词： Dog Alkaline phosphatase isoenzyme Glucocorticoid hepatopathy Ultrasonography

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Chinese Journal of Oceanology and Limnology 2014年第6期32卷 1364-1372页

作者：赵吉强李丽霞 College of Life Sciences Yantai University

Macroalgae in a littoral zone are inevitably exposed to UV-B irradiance. We analyzed the effects of UV-B on isoenzyme patterns and activities of superoxide dismutase （SOD）, peroxidase （POX）, catalase （CAT）, and ascorbate peroxidase （APX） of red algae Grateloupia filicina （Lamour.） C. Agardh. The activities of SOD, CAT, and APX changed in response to UV-B in a time- and dose-dependent manner. POX activity increased significantly under all three UV-B treatments. The enzymatic assay showed three distinct bands of SODI （Mn-SOD）, SODII （Fe-SOD）, and SODIII （CuZn-SOD） under a low （Luv） and medium （Muv） dose of UV-B irradiation, while SODI and SODIII activities decreased significantly when exposed to a high dose of UV-B irradiation （Huv）. The activity of POX isoenzymes increased significantly after exposure to UV-B, which is consistent with the total activity. In addition, a clear decrease in activity of CATIV was detected in response to all the three doses of UV treatments. Some bands ofAPX isoenzyme were also clearly influenced by UV-B irradiation. Correspondingly, the daily growth rate declined under all the three exposure doses, and was especially significant under Muv and Huv treatments. These data suggest that, although the protection mechanisms of antioxidant defense system are partly inducible by UV-B to prevent the damage, *** has incomplete tolerance to higher UV-B irradiation stress.

关键词： UV-B irradiation Grateloupiafilicina antioxidant system isoenzyme electrophoresis

Enhancement of Superoxide Dismutase and Catalase Activities and Salt Tolerance of Euhalophyte Suaeda salsa L. by Mycorrhizal Fungus Glomus mosseae

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Pedosphere 2012年第2期22卷 217-224页

作者： LI Tao LIU Run-Jin HE Xin-Hua WANG Bao-Shan Key Laboratory of Plant Stress Research College of Life SciencesShandong Normal UniversityJinan 250014(China) Institute of Mycorrhizal Biotechnology Qingdao Agricultural UniversityQingdao 266109(China) School of Plant Biology University of Western AustraliaCrawleyWA 6009(Australia) Northern Research Station USDA Forest Service and School of Forest Resources and Environmental ScienceMichigan Technological UniversityHoughtonMI 49931(USA)

Arbuscular mycorrhizal （AM）-mediated plant physiological activities could contribute to plant salt tolerance. However, the biochemical mechanism by which AM fungi enhance salt tolerance of halophytie plants is unclear. A pot experiment was conducted to determine whether salt tolerance of the C3 halophyte Suaeda salsa was enhanced by the AM fungus Glomus rnosseae. When 60-day-old S. salsa seedlings were subjected to 400 mmol L-1 NaC1 stress for 35 days, plant height, number of leaves and branches, shoot and root biomass, and root length of G. mosseae-colonized seedlings were significantly greater than those of the nonmycorrizal seedlings. Leaf superoxide dismutase （SOD） activity at all sampling times （weekly for 35 days after salt stress was initiated） and leaf catalase （CAT） activity at 2 and 3 weeks after salt stress was initiated were also significantly enhanced in G. mosseae-colonized S. salsa seedlings, while the content of leaf malondialdehyde （MDA）, a product of membrane lipid peroxidation, was significantly reduced, indicating an alleviation of oxidative damage. The corresponding leaf isoenzymes of SOD （Fe-SOD, Cu/Zn-SOD1, and Cu/Zn-SOD2） and CAT （CAT1 and CAT2） were also significantly increased in the mycorrhizal seedlings after 14 days of 400 mmol L-1 NaC1 stress. Our results suggested that G. rnosseae increased salt tolerance by increasing SOD and CAT activities and forming SOD and CAT isoforms in S. salsa seedlings.

关键词： antioxidant enzymes isoenzyme malondialdehyde NaC1 tolerance oxidative stress

Possible genetic reproductive isolation between two tilapiine genera and species:Oreochromis niloticus and Sarotherodon melanotheron

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Zoological Research 2011年第5期32卷 521-527页

作者：李思发赵岩范武江蔡完其许映芳上海海洋大学农业部水产种质资源与利用重点开放实验室上海201306 中国水产科学研究院长江水产研究所湖北荆州434000

Successful crossbreeding between Oreochromis niloticus and Sarotherodon melanotheron to produce a commercial hybrid has been *** karyotypes and isoenzyme of these two species and their reciprocal hybrids（*** ♀ × *** ♂,*** ♀ × *** ♂,the last not included in the isoenzyme study） were investigated via metaphase chromosomes obtained from head kidney cells and electropherogram of lactate dehydrogenase（LDH） isoenzymes from the liver,kidney,white muscle,heart,and eye *** diploid chromosome number（2n=44） and the fundamental number（NF=50） of the four tilapia genotypes were the ***,the karyotype of *** had three pairs of sub-metacentric（sm）,twelve pairs of sub-telocentric（st）,and seven pairs of telocentric（t） chromosomes,while *** had one pair of metacentric（m）,two pairs of sm,12 pairs of st,and seven pairs of t *** reciprocal hybrids both showed a mixed karyotype range between their parents：0.5 pair of m,2.5 pairs of sm,12 pairs of st,and seven pairs of t *** view of the electropherogram of isozymes,only the LDH of the kidney showed significant clear bands,with five bands in ***,three bands in ***,and duplicated six bands in the *** bands varied depending on their activities and *** considered that the differences in karyotype and isoenzyme were related to the genetic mechanism for post-mating isolation,and provided some additional basic genetic background of their taxonomy.

关键词： Tilapiine Oreochromis niloticus Sarotherodon melanotheron Karyotype isoenzyme Reproductive isolation Taxonomy