酶研究进展(英文)最新文献_第8页

Characterization of a Thermostable, Recombinant Carboxylesterase from the Hyperthermophilic Archaeon Metallosphaera sedula DSM5348 超嗜热古菌Metallosphaera sedula DSM5348的耐热重组羧酸酯酶的研究

酶研究进展(英文) Pub Date : 2014-03-19 DOI: 10.4236/AER.2014.21001

Rushyannah R. Killens-Cade, R. Turner, C. Macinnes, A. Grunden

{"title":"Characterization of a Thermostable, Recombinant Carboxylesterase from the Hyperthermophilic Archaeon Metallosphaera sedula DSM5348","authors":"Rushyannah R. Killens-Cade, R. Turner, C. Macinnes, A. Grunden","doi":"10.4236/AER.2014.21001","DOIUrl":"https://doi.org/10.4236/AER.2014.21001","url":null,"abstract":"Lipid-producing microalgae are emerging as the leading platform for producing alternative biofuels in response to diminishing petroleum reserves. Optimization of fatty acid production is required for efficient conversion of microalgal fatty acids into usable transportation fuels. Microbial lipases/esterases can be used to enhance fatty acid production because of their efficacy in catalyzing hydrolysis of esters into alcohols and fatty acids while minimizing the potential poisoning of catalysts needed in the biofuel production process. Although studies have extensively focused on lipases/esterases produced by mesophilic organisms, an understanding of lipases/esterases produced by thermophilic, acidic tolerant microbes, such as Metallosphaera sedula, is limited. In this work, the carboxylesterase from Metallosphaera sedula DSM5348 encoded by Msed_1072 was recombinantly expressed in Escherichia coli strain BL21 (λDE3). The purified enzyme either with a hexahistidine (His6)-tag (Msed_1072Nt and Msed_1072Ct) or without the hexahistidine (His6)-tag (Msed_1072) was biochemically characterized using a variety of substrates over a range of temperatures and pH and in the presence of metal ions, organic solvents, and detergents. In this study, the fusion of the protein with a hexahistidine (His6)-tag did not result in a change in substrate specificity, but the findings provide information on which enzyme variant can hydrolyze fatty acid esters in the presence of various chemicals, and this has important implication for their use in industrial processes. It also demonstrates that Metallosphaera sedula Msed_1072 can have application in microalgae-based biofuel production systems.","PeriodicalId":65616,"journal":{"name":"酶研究进展(英文)","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"70485025","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 5

Cloning and Expression of a Novel Phytase Gene (phyMS) from Mycobacterium smegmatis 耻垢分枝杆菌植酸酶基因的克隆与表达

酶研究进展(英文) Pub Date : 2014-03-19 DOI: 10.4236/AER.2014.21003

Tamrin Nuge, Y. Hashim, A. Farouk, H. Salleh

引用次数: 10

Papain Activity in Dextran Solution for Keratin Hydrolysis 葡聚糖水解角蛋白溶液中的木瓜蛋白酶活性

酶研究进展(英文) Pub Date : 2014-03-19 DOI: 10.4236/AER.2014.21005

Nanami Hara, Shintaro Morisada, K. Ohto, H. Kawakita

引用次数: 4

Parameter Estimation in Different Enzyme Reactions 不同酶反应的参数估计

酶研究进展(英文) Pub Date : 2014-03-19 DOI: 10.4236/AER.2014.21002

S. Nelatury, C. Nelatury, Mary Vagula

引用次数: 7

Purification and Characterisation of Xanthine Oxidoreductases from Local Bovids in Malta 马耳他当地牛科动物黄嘌呤氧化还原酶的纯化和鉴定

酶研究进展(英文) Pub Date : 2014-03-19 DOI: 10.4236/AER.2014.21006

M. Vella, T. Hunter, C. Farrugia, A. Pearson, G. Hunter

{"title":"Purification and Characterisation of Xanthine Oxidoreductases from Local Bovids in Malta","authors":"M. Vella, T. Hunter, C. Farrugia, A. Pearson, G. Hunter","doi":"10.4236/AER.2014.21006","DOIUrl":"https://doi.org/10.4236/AER.2014.21006","url":null,"abstract":"Xanthine oxidoreductase (XOR) is a molybdoflavoprotein mainly involved in purine catabolism. It exists in two forms, the oxidase (XO) and dehydrogenase (XDH) which are inter-convertible within mammalian cells. Although various researchers have reported the extraction of mammalian XOR, no extractions have yet been carried out in Malta and subsequently no characterizations are available. In this study, XOR was successfully purified from bovine, caprine and ovine milk through a multistep purification process involving both chemical and chromatographic techniques. The molecular weights of the native enzyme were found to be 295 kDa, 281 kDa and 275 kDa, representing the bovine, caprine and ovine XOR respectively. Western blot showed XOR to be represented on SDS-PAGE by a minimum of three major bands having molecular weights of 151 kDa, 131 kDa and 85 kDa. While all samples showed activity on native PAGE, spectrophotometric assays revealed the bovine XOR to be the most active. Surprisingly, the addition of NAD+ to the assay mixture inhibited enzyme activity of the bovine and caprine XOR whereas the ovine XOR doubled its activity in response to NAD+. The latter also showed a lower binding affinity to heparin. Following incubation with trypsin, XOR was irreversibly converted to its oxidase form in all samples as reflected by the observed increase in XO activity.","PeriodicalId":65616,"journal":{"name":"酶研究进展(英文)","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"70485499","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 4

Comparison of Substrate Specificity of Escherichia Coli p-Aminobenzoyl-Glutamate Hydrolase with Pseudomonas Carboxypeptidase G 大肠杆菌对氨基苯甲酰谷氨酸水解酶与假单胞菌羧肽酶G底物特异性的比较

酶研究进展(英文) Pub Date : 2014-03-01 DOI: 10.4236/aer.2014.21004

Cassandra M. Larimer, Dejan Slavnic, L. Pitstick, Jacalyn M. Green

{"title":"Comparison of Substrate Specificity of Escherichia Coli p-Aminobenzoyl-Glutamate Hydrolase with Pseudomonas Carboxypeptidase G","authors":"Cassandra M. Larimer, Dejan Slavnic, L. Pitstick, Jacalyn M. Green","doi":"10.4236/aer.2014.21004","DOIUrl":"https://doi.org/10.4236/aer.2014.21004","url":null,"abstract":"Reduced folic acid derivatives support biosynthesis of DNA, RNA and amino acids in bacteria as well as in eukaryotes, including humans. While the genes and steps for bacterial folic acid synthesis are known, those associated with folic acid catabolism are not well understood. A folate catabolite found in both humans and bacteria is p-aminobenzoyl-glutamate (PABA-GLU). The enzyme p-aminobenzoyl-glutamate hydrolase (PGH) breaks down PABA-GLU and is part of an apparent operon, the abg region, in E. coli. The subunits of PGH possess sequence and catalytic similarities to carboxypeptidase enzymes from Pseudomonas species. A comparison of the subunit sequences and activity of PGH, relative to carboxypeptidase enzymes, may lead to a better understanding of bacterial physiology and pathway evolution. We first compared the amino acid sequences of AbgA, AbgB and carboxypeptidase G2 from Pseudomonas sp. RS-16, which has been crystallized. Then we compared the enzyme activities of E. coli PGH and commercially available Pseudomonas carboxypeptidase G using spectrophotometric assays measuring cleavage of PABA-GLU, folate, aminopterin, methotrexate, 5-formyltetrahydrofolate, and 5-methyltetrahydrofolate. The Km and Vmax values for the folate and anti-folate substrates of PGH could not be determined, because the instrument reached its limit before the enzyme was saturated. Therefore, activity of PGH was compared to the activity of CPG, or normalized to PABA-GLU (nmole/min/µg). Relative to its activity with 10 µM PABA-GLU (100%), PGH cleaved glutamate from methotrexate (48%), aminopterin (45%) and folate (9%). Reduced folates leucovorin (5-formyltetrahydrofolate) and 5-methyltetrahydrofolate were not cleaved by PGH. Our data suggest that E. coli PGH is specific for PABA-GLU as its activity with natural folates (folate, 5-methyltetrahydrofolate, and leucovorin) was very poor. It does, however, have some ability to cleave anti-folates which may have clinical applications in treatment of chemotherapy overdose.","PeriodicalId":65616,"journal":{"name":"酶研究进展(英文)","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2014-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"70485478","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 8

Escherichia coli superoxide dismutase expression does not change in response to iron challenge during lag phase: Is the ferric uptake regulator to blame? 在迟滞期，大肠杆菌超氧化物歧化酶的表达不改变对铁的响应:铁摄取调节剂是罪魁祸首吗?

酶研究进展(英文) Pub Date : 2013-12-09 DOI: 10.4236/AER.2013.14014

R. Bertrand, M. Eze

{"title":"Escherichia coli superoxide dismutase expression does not change in response to iron challenge during lag phase: Is the ferric uptake regulator to blame?","authors":"R. Bertrand, M. Eze","doi":"10.4236/AER.2013.14014","DOIUrl":"https://doi.org/10.4236/AER.2013.14014","url":null,"abstract":"Intracellular iron levels and the expression of superoxide dismutase (SOD) and hydroperoxidase (HP) are regulated in Gram-negative bacteria by the iron(II)-activated ferric uptake regula- tor (Fur). We have previously observed that the expression of SOD in exponential phase Escherichia coli is dependent upon the redox state of iron in media, consistent with the ferrous specificity of Fur regulation (Bertrand et al., Med. Hypotheses 78: 130 - 133, 2012). Through the non-denaturing electrophoretic technique we have determined the Escherichia coli expression profiles of SOD and HP in response to iron challenge throughout lag, logarithmic, and stationary phases of replication. Lag phase SOD presented an unusual expression profile such that SOD expression was unresponsive to iron challenge, analogous to observations of mutant strains lacking Fur and of E. coli incubated in iron-deplete media. Challenging Escherichia coli with iron during logarithmic phase revealed that length of exposure to oxidants is unlikely to be the cause of SOD unresponsiveness in lag phase. HP activity was up-regulated two- or three-fold throughout all growth phases in response to iron challenge, but did not present redox- or growth phase-specific outcomes in a manner analogous to SOD. We hypothesize that low Fur levels during lag phase are responsible for unresponsive SOD.","PeriodicalId":65616,"journal":{"name":"酶研究进展(英文)","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2013-12-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"70484986","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 4

Optimized production and properties of thermostable alkaline protease from Bacillus subtilis SHS-04 grown on groundnut (Arachis hypogaea) meal 花生粕上生长的枯草芽孢杆菌SHS-04耐热碱性蛋白酶的优化生产及性能研究

酶研究进展(英文) Pub Date : 2013-12-09 DOI: 10.4236/AER.2013.14012

F. M. Olajuyigbe

引用次数: 17

Spinach aldolase interactions with rabbit, chicken, and fish muscle phosphofructokinase-1 菠菜醛缩酶与兔、鸡和鱼肌肉磷酸果糖激酶-1的相互作用

酶研究进展(英文) Pub Date : 2013-12-09 DOI: 10.4236/AER.2013.14013

Anita Williams, Ami Abbott, Jessica Chadwick, Alicia N. Thomas, Nathalia Cruz, A. Deng, Leah Ordinanza, John Tat, P. Russell

引用次数: 1

Partial purification, immobilization and preliminary biochemical characterization of lipases from Rhizomucor pusillus 根霉脂肪酶的部分纯化、固定化及初步生化特性研究

酶研究进展(英文) Pub Date : 2013-12-09 DOI: 10.4236/AER.2013.14009

A. L. Ferrarezi, D. Pivetta, G. O. Bonilla-Rodriguez, R. Silva, J. Guisán, E. Gomes, B. Pessela

{"title":"Partial purification, immobilization and preliminary biochemical characterization of lipases from Rhizomucor pusillus","authors":"A. L. Ferrarezi, D. Pivetta, G. O. Bonilla-Rodriguez, R. Silva, J. Guisán, E. Gomes, B. Pessela","doi":"10.4236/AER.2013.14009","DOIUrl":"https://doi.org/10.4236/AER.2013.14009","url":null,"abstract":"Lipases have important applications in biotechnological processes, motivating us to produce, purify, immobilize and perform a biochemical characterization of the lipase from Rhizomucor pusillus. The fungus was cultivated by solid state fermentation producing lipolytic activity of about 0.5 U/mL(4U/g). A partial purification by gel filtration chromatography in Se-phacryl S-100 allowed obtaining a yield of about 85% and a purification factor of 5.7. Our results revealed that the purified enzyme is very stable with some significant differences in its properties when compared to crude extract. The crude enzyme extract has an optimum pH and temperature of 7.5 ° C and 40 ° C, respectively. After purification, a shift of the optimum pH from 7 to 8 was observed, as well as a rise in optimumtemperature to 60 ° C and an increase in stability. The enzyme was immobilized on CNBr-Agarose and Octyl-Agarose supports, having the highest immobilization yield of 94% in the second resin. The major advantage of immobilization in hydrophobic media such as Octyl is in its hyper activation, which in this case was over 200%, a very interesting finding. Another advantage of this type of immobilization is the possibility of using the derivatives in biotechnological applications, such as in oil enriched with omega-3 as the results obtained in this study display the hydrolysis of 40% EPA and 7% DHA from sardine oil, promising results compared to the literature.","PeriodicalId":65616,"journal":{"name":"酶研究进展(英文)","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2013-12-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"70484686","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 4