Hyperexpression of a Bacillus circulans xylanase gene in Escherichia coli and characterization of the gene product
A 4.0-kilobase (kb) fragment of Bacillus circulans genomic DNA inserted into pUC19 and encoding endoxylanase activity was subjected to a series of subclonings. A 1.0-kb HindIII-HincII subfragment was found to code for xylanase activity. Maximum expression levels were observed with a subclone that co...
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| Veröffentlicht in: | Applied and Environmental Microbiology 1989-05, Vol.55 (5), p.1192-1195 |
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| creator | Yang, R.C.A MacKenzie, C.R Bilous, D Narang, S.A |
| description | A 4.0-kilobase (kb) fragment of Bacillus circulans genomic DNA inserted into pUC19 and encoding endoxylanase activity was subjected to a series of subclonings. A 1.0-kb HindIII-HincII subfragment was found to code for xylanase activity. Maximum expression levels were observed with a subclone that contained an additional 0.3-kb sequence upstream from the coding region. Enhancer sequences in the upstream region are thought to be responsible for these high expression levels. Southern hybridization analyses revealed that the cloned gene hybridized with genomic DNA from Bacillus subtilis and Bacillus polymyxa. Xylanase activity expressed by Escherichia coli harboring the cloned gene was located primarily in the intracellular fraction.Levels of up to 7 U/ml or 35 mg/liter were obtained. The protein product was purified by ion exchange and gel permeation chromatography. The xylanase had a molecular weight of 20,500 and an isoelectric point of 9.0 |
| doi_str_mv | 10.1128/aem.55.5.1192-1195.1989 |
| format | Article |
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A 1.0-kb HindIII-HincII subfragment was found to code for xylanase activity. Maximum expression levels were observed with a subclone that contained an additional 0.3-kb sequence upstream from the coding region. Enhancer sequences in the upstream region are thought to be responsible for these high expression levels. Southern hybridization analyses revealed that the cloned gene hybridized with genomic DNA from Bacillus subtilis and Bacillus polymyxa. Xylanase activity expressed by Escherichia coli harboring the cloned gene was located primarily in the intracellular fraction.Levels of up to 7 U/ml or 35 mg/liter were obtained. The protein product was purified by ion exchange and gel permeation chromatography. The xylanase had a molecular weight of 20,500 and an isoelectric point of 9.0</description><identifier>ISSN: 0099-2240</identifier><identifier>EISSN: 1098-5336</identifier><identifier>DOI: 10.1128/aem.55.5.1192-1195.1989</identifier><identifier>PMID: 2667461</identifier><identifier>CODEN: AEMIDF</identifier><language>eng</language><publisher>Washington, DC: American Society for Microbiology</publisher><subject>ACTIVIDAD ENZIMATICA ; ACTIVITE ENZYMATIQUE ; AZUCARES ; BACILLUS ; Bacillus - enzymology ; Bacillus - genetics ; Biological and medical sciences ; Blotting, Southern ; Chromatography, Gel ; Chromatography, Ion Exchange ; CLONE ; CLONES ; Cloning, Molecular ; DNA, Bacterial - genetics ; Electrophoresis, Polyacrylamide Gel ; ENZIMAS ; ENZYME ; ESCHERICHIA ; Escherichia coli ; Escherichia coli - genetics ; Fundamental and applied biological sciences. Psychology ; GENE ; Gene expression ; Gene Expression Regulation ; gene products ; GENES ; Glycoside Hydrolases - biosynthesis ; Glycoside Hydrolases - genetics ; Glycoside Hydrolases - isolation & purification ; Molecular and cellular biology ; Molecular genetics ; Nucleic Acid Hybridization ; PARED CELULAR ; PAROI CELLULAIRE ; PLANTAS ; PLANTE ; Restriction Mapping ; SUCRES ; Xylan Endo-1,3-beta-Xylosidase</subject><ispartof>Applied and Environmental Microbiology, 1989-05, Vol.55 (5), p.1192-1195</ispartof><rights>1990 INIST-CNRS</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c606t-828da455f5652dff7a589069d26b5742a9870a1d80991f60feb72991ed85190c3</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC184275/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC184275/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,314,727,780,784,885,3188,3189,27924,27925,53791,53793</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=6743243$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/2667461$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Yang, R.C.A</creatorcontrib><creatorcontrib>MacKenzie, C.R</creatorcontrib><creatorcontrib>Bilous, D</creatorcontrib><creatorcontrib>Narang, S.A</creatorcontrib><title>Hyperexpression of a Bacillus circulans xylanase gene in Escherichia coli and characterization of the gene product</title><title>Applied and Environmental Microbiology</title><addtitle>Appl Environ Microbiol</addtitle><description>A 4.0-kilobase (kb) fragment of Bacillus circulans genomic DNA inserted into pUC19 and encoding endoxylanase activity was subjected to a series of subclonings. A 1.0-kb HindIII-HincII subfragment was found to code for xylanase activity. Maximum expression levels were observed with a subclone that contained an additional 0.3-kb sequence upstream from the coding region. Enhancer sequences in the upstream region are thought to be responsible for these high expression levels. Southern hybridization analyses revealed that the cloned gene hybridized with genomic DNA from Bacillus subtilis and Bacillus polymyxa. Xylanase activity expressed by Escherichia coli harboring the cloned gene was located primarily in the intracellular fraction.Levels of up to 7 U/ml or 35 mg/liter were obtained. The protein product was purified by ion exchange and gel permeation chromatography. The xylanase had a molecular weight of 20,500 and an isoelectric point of 9.0</description><subject>ACTIVIDAD ENZIMATICA</subject><subject>ACTIVITE ENZYMATIQUE</subject><subject>AZUCARES</subject><subject>BACILLUS</subject><subject>Bacillus - enzymology</subject><subject>Bacillus - genetics</subject><subject>Biological and medical sciences</subject><subject>Blotting, Southern</subject><subject>Chromatography, Gel</subject><subject>Chromatography, Ion Exchange</subject><subject>CLONE</subject><subject>CLONES</subject><subject>Cloning, Molecular</subject><subject>DNA, Bacterial - genetics</subject><subject>Electrophoresis, Polyacrylamide Gel</subject><subject>ENZIMAS</subject><subject>ENZYME</subject><subject>ESCHERICHIA</subject><subject>Escherichia coli</subject><subject>Escherichia coli - genetics</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>GENE</subject><subject>Gene expression</subject><subject>Gene Expression Regulation</subject><subject>gene products</subject><subject>GENES</subject><subject>Glycoside Hydrolases - biosynthesis</subject><subject>Glycoside Hydrolases - genetics</subject><subject>Glycoside Hydrolases - isolation & purification</subject><subject>Molecular and cellular biology</subject><subject>Molecular genetics</subject><subject>Nucleic Acid Hybridization</subject><subject>PARED CELULAR</subject><subject>PAROI CELLULAIRE</subject><subject>PLANTAS</subject><subject>PLANTE</subject><subject>Restriction Mapping</subject><subject>SUCRES</subject><subject>Xylan Endo-1,3-beta-Xylosidase</subject><issn>0099-2240</issn><issn>1098-5336</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1989</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkU2P0zAQhi0EWkrhDyAhjIS4pdhO_HXYA6wWFmklDrBna-rYjVESFzuBLb8eV40q9sTFM_Y879jjF6HXlGwoZeo9uGHD-YaXnWZVWUqmlX6EVpRoVfG6Fo_RihCtK8Ya8hQ9y_kHIaQhQl2gCyaEbARdoXRz2Lvk7vfJ5RziiKPHgD-CDX0_Z2xDsnMPY8b3hxIgO7xzo8NhxNfZdi4F2wXANvYBw9hi20ECO5XzPzAt7aZuEe1TbGc7PUdPPPTZvVjiGt19uv5-dVPdfv385erDbWUFEVOlmGqh4dxzwVnrvQSuNBG6ZWLLZcNAK0mAtqrMSL0g3m0lK6lrFaea2HqNLk999_N2cK1145SgN_sUBkgHEyGYh5UxdGYXfxmqGiZ50b9b9Cn-nF2ezBCydX35BxfnbKSmtajJ_0HKmZSk0GskT6BNMefk_PkxlJijrabYajg33BxtPS4lK7YW5at_ZznrFh9L_e1Sh2yh9wlGG_IZK1DNmrpgb05YF3bd75CcgTw8vLQwL0-Mh2hgl0qbu29KMyGVqv8CtTDBSQ</recordid><startdate>19890501</startdate><enddate>19890501</enddate><creator>Yang, R.C.A</creator><creator>MacKenzie, C.R</creator><creator>Bilous, D</creator><creator>Narang, S.A</creator><general>American Society for Microbiology</general><scope>FBQ</scope><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QL</scope><scope>7QO</scope><scope>7TM</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>19890501</creationdate><title>Hyperexpression of a Bacillus circulans xylanase gene in Escherichia coli and characterization of the gene product</title><author>Yang, R.C.A ; MacKenzie, C.R ; Bilous, D ; Narang, S.A</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c606t-828da455f5652dff7a589069d26b5742a9870a1d80991f60feb72991ed85190c3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1989</creationdate><topic>ACTIVIDAD ENZIMATICA</topic><topic>ACTIVITE ENZYMATIQUE</topic><topic>AZUCARES</topic><topic>BACILLUS</topic><topic>Bacillus - enzymology</topic><topic>Bacillus - genetics</topic><topic>Biological and medical sciences</topic><topic>Blotting, Southern</topic><topic>Chromatography, Gel</topic><topic>Chromatography, Ion Exchange</topic><topic>CLONE</topic><topic>CLONES</topic><topic>Cloning, Molecular</topic><topic>DNA, Bacterial - genetics</topic><topic>Electrophoresis, Polyacrylamide Gel</topic><topic>ENZIMAS</topic><topic>ENZYME</topic><topic>ESCHERICHIA</topic><topic>Escherichia coli</topic><topic>Escherichia coli - genetics</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>GENE</topic><topic>Gene expression</topic><topic>Gene Expression Regulation</topic><topic>gene products</topic><topic>GENES</topic><topic>Glycoside Hydrolases - biosynthesis</topic><topic>Glycoside Hydrolases - genetics</topic><topic>Glycoside Hydrolases - isolation & purification</topic><topic>Molecular and cellular biology</topic><topic>Molecular genetics</topic><topic>Nucleic Acid Hybridization</topic><topic>PARED CELULAR</topic><topic>PAROI CELLULAIRE</topic><topic>PLANTAS</topic><topic>PLANTE</topic><topic>Restriction Mapping</topic><topic>SUCRES</topic><topic>Xylan Endo-1,3-beta-Xylosidase</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Yang, R.C.A</creatorcontrib><creatorcontrib>MacKenzie, C.R</creatorcontrib><creatorcontrib>Bilous, D</creatorcontrib><creatorcontrib>Narang, S.A</creatorcontrib><collection>AGRIS</collection><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Biotechnology Research Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Applied and Environmental Microbiology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Yang, R.C.A</au><au>MacKenzie, C.R</au><au>Bilous, D</au><au>Narang, S.A</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Hyperexpression of a Bacillus circulans xylanase gene in Escherichia coli and characterization of the gene product</atitle><jtitle>Applied and Environmental Microbiology</jtitle><addtitle>Appl Environ Microbiol</addtitle><date>1989-05-01</date><risdate>1989</risdate><volume>55</volume><issue>5</issue><spage>1192</spage><epage>1195</epage><pages>1192-1195</pages><issn>0099-2240</issn><eissn>1098-5336</eissn><coden>AEMIDF</coden><abstract>A 4.0-kilobase (kb) fragment of Bacillus circulans genomic DNA inserted into pUC19 and encoding endoxylanase activity was subjected to a series of subclonings. A 1.0-kb HindIII-HincII subfragment was found to code for xylanase activity. Maximum expression levels were observed with a subclone that contained an additional 0.3-kb sequence upstream from the coding region. Enhancer sequences in the upstream region are thought to be responsible for these high expression levels. Southern hybridization analyses revealed that the cloned gene hybridized with genomic DNA from Bacillus subtilis and Bacillus polymyxa. Xylanase activity expressed by Escherichia coli harboring the cloned gene was located primarily in the intracellular fraction.Levels of up to 7 U/ml or 35 mg/liter were obtained. The protein product was purified by ion exchange and gel permeation chromatography. The xylanase had a molecular weight of 20,500 and an isoelectric point of 9.0</abstract><cop>Washington, DC</cop><pub>American Society for Microbiology</pub><pmid>2667461</pmid><doi>10.1128/aem.55.5.1192-1195.1989</doi><tpages>4</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | Applied and Environmental Microbiology, 1989-05, Vol.55 (5), p.1192-1195 |
| issn | 0099-2240 1098-5336 |
| language | eng |
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| source | American Society for Microbiology; MEDLINE; PubMed Central; Alma/SFX Local Collection |
| subjects | ACTIVIDAD ENZIMATICA ACTIVITE ENZYMATIQUE AZUCARES BACILLUS Bacillus - enzymology Bacillus - genetics Biological and medical sciences Blotting, Southern Chromatography, Gel Chromatography, Ion Exchange CLONE CLONES Cloning, Molecular DNA, Bacterial - genetics Electrophoresis, Polyacrylamide Gel ENZIMAS ENZYME ESCHERICHIA Escherichia coli Escherichia coli - genetics Fundamental and applied biological sciences. Psychology GENE Gene expression Gene Expression Regulation gene products GENES Glycoside Hydrolases - biosynthesis Glycoside Hydrolases - genetics Glycoside Hydrolases - isolation & purification Molecular and cellular biology Molecular genetics Nucleic Acid Hybridization PARED CELULAR PAROI CELLULAIRE PLANTAS PLANTE Restriction Mapping SUCRES Xylan Endo-1,3-beta-Xylosidase |
| title | Hyperexpression of a Bacillus circulans xylanase gene in Escherichia coli and characterization of the gene product |
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