Enzyme Research最新文献_第5页

Long-Range PCR Amplification of DNA by DNA Polymerase III Holoenzyme from Thermus thermophilus. 嗜热热菌DNA聚合酶III全酶的远程PCR扩增。

Enzyme Research Pub Date : 2015-01-01 Epub Date: 2015-01-19 DOI: 10.1155/2015/837842

Wendy Ribble, Shawn D Kane, James M Bullard

{"title":"Long-Range PCR Amplification of DNA by DNA Polymerase III Holoenzyme from Thermus thermophilus.","authors":"Wendy Ribble, Shawn D Kane, James M Bullard","doi":"10.1155/2015/837842","DOIUrl":"https://doi.org/10.1155/2015/837842","url":null,"abstract":"DNA replication in bacteria is accomplished by a multicomponent replicase, the DNA polymerase III holoenzyme (pol III HE). The three essential components of the pol III HE are the α polymerase, the β sliding clamp processivity factor, and the DnaX clamp-loader complex. We report here the assembly of the functional holoenzyme from Thermus thermophilus (Tth), an extreme thermophile. The minimal holoenzyme capable of DNA synthesis consists of α, β and DnaX (τ and γ), δ and δ' components of the clamp-loader complex. The proteins were each cloned and expressed in a native form. Each component of the system was purified extensively. The minimum holoenzyme from these five purified subunits reassembled is sufficient for rapid and processive DNA synthesis. In an isolated form the α polymerase was found to be unstable at temperatures above 65°C. We were able to increase the thermostability of the pol III HE to 98°C by addition and optimization of various buffers and cosolvents. In the optimized buffer system we show that a replicative polymerase apparatus, Tth pol III HE, is capable of rapid amplification of regions of DNA up to 15,000 base pairs in PCR reactions. ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2015-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1155/2015/837842","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33060976","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 4

Actinomycetes: A Source of Lignocellulolytic Enzymes. 放线菌：木质纤维分解酶的来源

Enzyme Research Pub Date : 2015-01-01 Epub Date: 2015-12-17 DOI: 10.1155/2015/279381

Anita Saini, Neeraj K Aggarwal, Anuja Sharma, Anita Yadav

{"title":"Actinomycetes: A Source of Lignocellulolytic Enzymes.","authors":"Anita Saini, Neeraj K Aggarwal, Anuja Sharma, Anita Yadav","doi":"10.1155/2015/279381","DOIUrl":"10.1155/2015/279381","url":null,"abstract":"Lignocellulose is the most abundant biomass on earth. Agricultural, forest, and agroindustrial activities generate tons of lignocellulosic wastes annually, which present readily procurable, economically affordable, and renewable feedstock for various lignocelluloses based applications. Lignocelluloses are the focus of present decade researchers globally, in an attempt to develop technologies based on natural biomass for reducing dependence on expensive and exhaustible substrates. Lignocellulolytic enzymes, that is, cellulases, hemicellulases, and lignolytic enzymes, play very important role in the processing of lignocelluloses which is prerequisite for their utilization in various processes. These enzymes are obtained from microorganisms distributed in both prokaryotic and eukaryotic domains including bacteria, fungi, and actinomycetes. Actinomycetes are an attractive microbial group for production of lignocellulose degrading enzymes. Various studies have evaluated the lignocellulose degrading ability of actinomycetes, which can be potentially implemented in the production of different value added products. This paper is an overview of the diversity of cellulolytic, hemicellulolytic, and lignolytic actinomycetes along with brief discussion of their hydrolytic enzyme systems involved in biomass modification. ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2015-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4697097/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"86269036","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Optimization of Enzymatic Saccharification of Alkali Pretreated Parthenium sp. Using Response Surface Methodology. 利用响应面方法优化碱处理帕氏藻的酶糖化过程

Enzyme Research Pub Date : 2014-01-01 Epub Date: 2014-05-12 DOI: 10.1155/2014/764898

K Pandiyan, Rameshwar Tiwari, Surender Singh, Pawan K S Nain, Sarika Rana, Anju Arora, Shashi B Singh, Lata Nain

{"title":"Optimization of Enzymatic Saccharification of Alkali Pretreated Parthenium sp. Using Response Surface Methodology.","authors":"K Pandiyan, Rameshwar Tiwari, Surender Singh, Pawan K S Nain, Sarika Rana, Anju Arora, Shashi B Singh, Lata Nain","doi":"10.1155/2014/764898","DOIUrl":"10.1155/2014/764898","url":null,"abstract":"Parthenium sp. is a noxious weed which threatens the environment and biodiversity due to its rapid invasion. This lignocellulosic weed was investigated for its potential in biofuel production by subjecting it to mild alkali pretreatment followed by enzymatic saccharification which resulted in significant amount of fermentable sugar yield (76.6%). Optimization of enzymatic hydrolysis variables such as temperature, pH, enzyme, and substrate loading was carried out using central composite design (CCD) in response to surface methodology (RSM) to achieve the maximum saccharification yield. Data obtained from RSM was validated using ANOVA. After the optimization process, a model was proposed with predicted value of 80.08% saccharification yield under optimum conditions which was confirmed by the experimental value of 85.80%. This illustrated a good agreement between predicted and experimental response (saccharification yield). The saccharification yield was enhanced by enzyme loading and reduced by temperature and substrate loading. This study reveals that under optimized condition, sugar yield was significantly increased which was higher than earlier reports and promises the use of Parthenium sp. biomass as a feedstock for bioethanol production. ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4036719/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32400330","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Purification and characterization of melanogenic enzyme tyrosinase from button mushroom. 钮扣菇产黑素酶酪氨酸酶的纯化及特性研究。

Enzyme Research Pub Date : 2014-01-01 Epub Date: 2014-08-14 DOI: 10.1155/2014/120739

Kamal Uddin Zaidi, Ayesha S Ali, Sharique A Ali

{"title":"Purification and characterization of melanogenic enzyme tyrosinase from button mushroom.","authors":"Kamal Uddin Zaidi, Ayesha S Ali, Sharique A Ali","doi":"10.1155/2014/120739","DOIUrl":"https://doi.org/10.1155/2014/120739","url":null,"abstract":"Melanogenesis is a biosynthetic pathway for the formation of the pigment melanin in human skin. A key enzyme, tyrosinase, catalyzes the first and only rate-limiting steps in melanogenesis. Since the discovery of its melanogenic properties, tyrosinase has been in prime focus and microbial sources of the enzyme are sought. Agaricus bisporus widely known as the common edible mushroom, it's taking place in high amounts of proteins, enzyme, carbohydrates, fibers, and low fat contents are frequently cited in the literature in relation to their nutritional value. In the present study tyrosinase from Agaricus bisporus was purified by ammonium sulphate precipitation, dialysis followed by gel filtration chromatography on Sephadex G-100, and ion exchange chromatography on DEAE-Cellulose; the enzyme was purified, 16.36-fold to give 26.6% yield on total activity in the crude extract and final specific activity of 52.19 U/mg. The SDS-PAGE electrophoresis showed a migrating protein band molecular weight of 95 kDa. The purified tyrosinase was optimized and the results revealed that the optimum values are pH 7.0 and temperature 35°C. The highest activity was reported towards its natural substrate, L-DOPA, with an apparent Km value of 0.933 mM. This indicated that tyrosinase purified from Agaricus bisporus is a potential source for medical applications. ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1155/2014/120739","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32647428","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 45

Aspergillus 6V4, a Strain Isolated from Manipueira, Produces High Amylases Levels by Using Wheat Bran as a Substrate. 以麦麸为底物制备高淀粉酶的曲霉6V4菌株

Enzyme Research Pub Date : 2014-01-01 Epub Date: 2014-03-02 DOI: 10.1155/2014/725651

Jessyca Dos Reis Celestino, Ana Caroline Duarte, Cláudia Maria de Melo Silva, Hellen Holanda Sena, Maria do Perpétuo Socorro Borges Carriço Ferreira, Neila Hiraishi Mallmann, Natacha Pinheiro Costa Lima, Chanderlei de Castro Tavares, Rodrigo Otávio Silva de Souza, Erica Simplício Souza, João Vicente Braga Souza

{"title":"Aspergillus 6V4, a Strain Isolated from Manipueira, Produces High Amylases Levels by Using Wheat Bran as a Substrate.","authors":"Jessyca Dos Reis Celestino, Ana Caroline Duarte, Cláudia Maria de Melo Silva, Hellen Holanda Sena, Maria do Perpétuo Socorro Borges Carriço Ferreira, Neila Hiraishi Mallmann, Natacha Pinheiro Costa Lima, Chanderlei de Castro Tavares, Rodrigo Otávio Silva de Souza, Erica Simplício Souza, João Vicente Braga Souza","doi":"10.1155/2014/725651","DOIUrl":"https://doi.org/10.1155/2014/725651","url":null,"abstract":"The aim of this study was screening fungi strains, isolated from manipueira (a liquid subproduct obtained from the flour production of Manihot esculenta), for amylases production and investigating production of these enzymes by the strain Aspergillus 6V4. The fungi isolated from manipueira belonged to Ascomycota phylum. The strain Aspergillus 6V4 was the best amylase producer in the screening assay of starch hydrolysis in petri dishes (ASHPD) and in the assay in submerged fermentation (ASbF). The strain Aspergillus 6V4 produced high amylase levels (335 UI/L) using wheat bran infusion as the exclusive substrate and the supplementation of this substrate with peptone decreased the production of this enzyme. The moisture content of 70% was the best condition for the production of Aspergillus 6V4 amylases (385 IU/g) in solid state fermentation (SSF). ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1155/2014/725651","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32255327","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 5

Immobilization of a Plant Lipase from Pachira aquatica in Alginate and Alginate/PVA Beads. 海藻酸酯和海藻酸酯/聚乙烯醇微球固定化水生Pachira脂肪酶的研究。

Enzyme Research Pub Date : 2014-01-01 Epub Date: 2014-04-10 DOI: 10.1155/2014/738739

Bárbara M Bonine, Patricia Peres Polizelli, Gustavo O Bonilla-Rodriguez

引用次数: 80

A cDNA Cloning of a Novel Alpha-Class Tyrosinase of Pinctada fucata: Its Expression Analysis and Characterization of the Expressed Protein. 一种新型fucata α - class酪氨酸酶的cDNA克隆及其表达分析和表达蛋白的鉴定。

Enzyme Research Pub Date : 2014-01-01 Epub Date: 2014-04-10 DOI: 10.1155/2014/780549

Ryousuke Takgi, Tomoyuki Miyashita

{"title":"A cDNA Cloning of a Novel Alpha-Class Tyrosinase of Pinctada fucata: Its Expression Analysis and Characterization of the Expressed Protein.","authors":"Ryousuke Takgi, Tomoyuki Miyashita","doi":"10.1155/2014/780549","DOIUrl":"https://doi.org/10.1155/2014/780549","url":null,"abstract":"Tyrosinase plays an important role in the formation of the shell matrix and melanin synthesis in mollusks shells. A cDNA clone encoding a 47 kDa protein was isolated from the pearl oyster Pinctada fucata. The cDNA was 1,957 base pairs long and encodes a 417 residue protein that has extensive sequence identity with tyrosinase (polyphenol oxidase: EC 1.14.18.1). This tyrosinase-like protein, termed PfTy, contains an N-terminal signal sequence and the two copper-binding domain signatures (CuA and CuB), suggesting that PfTy belongs to the α -subclass of type-3 copper proteins. Enzyme activity of PfTy was examined by a spectrophotometric method using the translation product derived from an S30 T7 high-yield protein expression system. Tyrosinase activity was seen in this recombinant product. RT-PCR analysis showed that PfTy mRNA was expressed in the mantle pallial, but not in the mantle edge. Therefore, PfTy may participate in insoluble shell matrix formation of the nacreous layer. PfTy expression was also observed in the foot, liver, and adductor muscle, suggesting that PfTy participates in the synthesis of melanins, which are effective scavengers of free radicals formed in multiple intracellular oxidative processes. This is the first report of a novel α -class tyrosinase from the pearl oyster P. fucata. ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1155/2014/780549","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32333195","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 24

Effect of Temperature on Xylanase II from Trichoderma reesei QM 9414: A Calorimetric, Catalytic, and Conformational Study. 温度对里氏木霉qm9414木聚糖酶II的影响:量热、催化和构象研究。

Enzyme Research Pub Date : 2014-01-01 Epub Date: 2014-09-07 DOI: 10.1155/2014/708676

Gloria López, Pilar Estrada

{"title":"Effect of Temperature on Xylanase II from Trichoderma reesei QM 9414: A Calorimetric, Catalytic, and Conformational Study.","authors":"Gloria López, Pilar Estrada","doi":"10.1155/2014/708676","DOIUrl":"https://doi.org/10.1155/2014/708676","url":null,"abstract":"The secondary structure of xylanase II from Trichoderma reesei is lost in an apparent irreversible cooperative process as temperature is increased with a midpoint transition of 58.8 ± 0.1°C. The shift of the spectral centre of mass above 50°C is also apparently cooperative with midpoint transition of 56.3 ± 0.2°C, but the existence of two isofluorescent points in the fluorescence emission spectra suggests a non-two-state process. Further corroboration comes from differential scanning calorimetry experiments. At protein concentrations ≤0.56 mg·mL(-1) the calorimetric transition is reversible and the data were fitted to a non-two-state model and deconvoluted into six transitions, whereas at concentrations greater than 0.56 mg·mL(-1) the calorimetric transition is irreversible with an exothermic contribution to the thermogram. The apparent T m increased linearly with the scan rate according to first order inactivation kinetics. The effect of additives on the calorimetric transition of xylanase is dependent on their nature. The addition of sorbitol transforms reversible transitions into irreversible transitions while stabilizing the protein as the apparent T m increases linearly with sorbitol concentration. d-Glucono-1,5-lactone, a noncompetitive inhibitor in xylanase kinetics, and soluble xylan change irreversible processes into reversible processes at high protein concentration. ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1155/2014/708676","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32715131","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 9

Angiotensin converting enzyme activity in alopecia areata. 斑秃血管紧张素转换酶活性。

Enzyme Research Pub Date : 2014-01-01 Epub Date: 2014-10-01 DOI: 10.1155/2014/694148

Mohammad Reza Namazi, Armaghan Ashraf, Farhad Handjani, Ebrahim Eftekhar, Amir Kalafi

{"title":"Angiotensin converting enzyme activity in alopecia areata.","authors":"Mohammad Reza Namazi, Armaghan Ashraf, Farhad Handjani, Ebrahim Eftekhar, Amir Kalafi","doi":"10.1155/2014/694148","DOIUrl":"https://doi.org/10.1155/2014/694148","url":null,"abstract":"Background. Alopecia areata (AA) is a chronic inflammatory disease of the hair follicle. The exact pathogenesis of AA remains unknown, although recent studies support a T-cell mediated autoimmune process. On the other hand, some studies have proposed that the renin-angiotensin-aldosterone system (RAAS) may play a role in autoimmunity. Therefore, we assessed serum activity of angiotensin converting enzyme (ACE), a component of this system, in AA. Methods. ACE activity was measured in the sera of 19 patients with AA and 16 healthy control subjects. In addition, the relationship between severity and duration of the disease and ACE activity was evaluated. Results. Serum ACE activity was higher in the patient group (55.81 U/L) compared to the control group (46.41 U/L), but the difference was not statistically significant (P = 0.085). Also, there was no correlation between ACE activity and severity (P = 0.13) and duration of disease (P = 0.25) in the patient group. Conclusion. The increased serum ACE activity found in this study may demonstrate local involvement of the RAAS in the pathogenesis of AA. Assessment of ACE in a study with a larger sample size as well as in tissue samples is recommended in order to further evaluate the possible role of RAAS in AA. ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1155/2014/694148","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"32774782","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 6

Purification and characterization of glucose-6-phosphate dehydrogenase from camel liver. 从骆驼肝脏中纯化葡萄糖-6-磷酸脱氢酶并确定其特性。

Enzyme Research Pub Date : 2014-01-01 Epub Date: 2014-12-25 DOI: 10.1155/2014/714054

Mahmoud A Ibrahim, Abdel-Hady M Ghazy, Ahmed M H Salem, Mohamed A Ghazy, Mohamed M Abdel-Monsef

{"title":"Purification and characterization of glucose-6-phosphate dehydrogenase from camel liver.","authors":"Mahmoud A Ibrahim, Abdel-Hady M Ghazy, Ahmed M H Salem, Mohamed A Ghazy, Mohamed M Abdel-Monsef","doi":"10.1155/2014/714054","DOIUrl":"10.1155/2014/714054","url":null,"abstract":"Glucose-6-phosphate dehydrogenase from camel liver was purified to homogeneity by ammonium sulfate precipitation and a combination of DEAE-cellulose, Sephacryl S-300 gel filtration, and 2', 5' ADP Sepharose 4B affinity chromatography columns. The specific activity of camel liver G6PD is increased to 1.80438 units/mg proteins with 63-fold purification. It turned out to be homogenous on both native PAGE and 12% SDS PAGE, with a molecular weight of 64 kDa. The molecular weight of the native form of camel liver G6PD was determined to be 194 kDa by gel filtration indicating a trimeric protein. The K m value was found to be 0.081 mM of NADP(+). Camel liver G6PD displayed its optimum activity at pH 7.8 with an isoelectric point (pI) of pH 6.6-6.8. The divalent cations MgCl2, MnCl2, and CoCl2 act as activators; on the other hand, CaCl2 and NiCl2 act as moderate inhibitors, while FeCl2, CuCl2, and ZnCl2 are potent inhibitors of camel liver G6PD activity. NADPH inhibited camel liver G6PD competitively with K i value of 0.035 mM. One binding site was deduced for NADPH on the enzyme molecule. This study presents a simple and reproducible purification procedure of G6PD from the camel liver. ","PeriodicalId":11835,"journal":{"name":"Enzyme Research","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4290037/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33323621","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0