Expression and purification of soluble monomeric streptavidin in Escherichia coli
We recently reported the engineering of monomeric streptavidin (mSA) for use in monomeric detection of biotinylated ligands. Although mSA can be expressed functionally on the surface of mammalian cells and yeast, the molecule does not fold correctly when expressed in Escherichia coli. Refolding from...
Gespeichert in:
| Veröffentlicht in: | Applied microbiology and biotechnology 2014, Vol.98 (14), p.6285-6295 |
|---|---|
| Hauptverfasser: | , , |
| Format: | Artikel |
| Sprache: | eng |
| Schlagworte: | |
| Online-Zugang: | Volltext |
| Tags: |
Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
|
| container_end_page | 6295 |
|---|---|
| container_issue | 14 |
| container_start_page | 6285 |
| container_title | Applied microbiology and biotechnology |
| container_volume | 98 |
| creator | Demonte, Daniel Dundas, Christopher M Park, Sheldon |
| description | We recently reported the engineering of monomeric streptavidin (mSA) for use in monomeric detection of biotinylated ligands. Although mSA can be expressed functionally on the surface of mammalian cells and yeast, the molecule does not fold correctly when expressed in Escherichia coli. Refolding from inclusion bodies is cumbersome and yields a limited amount of purified protein. Improving the final yield should facilitate its use in biotechnology. We tested the expression and purification of mSA fused to GST, MBP, thioredoxin, and sumo tags to simplify its purification and improve the yield. The fusion proteins can be expressed solubly in E. coli and increase the yield by more than 20-fold. Unmodified mSA can be obtained by proteolytically removing the fusion tag. Purified mSA can be immobilized on a solid matrix to purify biotinylated ligands. Together, expressing mSA as a fusion with a solubilization tag vastly simplifies its preparation and increases its usability in biotechnology. |
| doi_str_mv | 10.1007/s00253-014-5682-y |
| format | Article |
| fullrecord | <record><control><sourceid>gale_proqu</sourceid><recordid>TN_cdi_proquest_miscellaneous_1554942714</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><galeid>A379569853</galeid><sourcerecordid>A379569853</sourcerecordid><originalsourceid>FETCH-LOGICAL-c637t-ea028a916d1ade7451f464cb793e8ab97975da1cea0703db44382b49c94acde23</originalsourceid><addsrcrecordid>eNqFkt1r1TAYxoMo7mz6B3ijBW_mRWe-01yOcdTBQHTuOqTp27OMtqlJKzv_vSmdH0dESSDwvr_ngSc8CL0g-IxgrN4mjKlgJSa8FLKi5f4R2hDOaIkl4Y_RBhMlSiV0dYSOU7rDmNBKyqfoiHKpSSXVBn3a3o8RUvJhKOzQFOMcfeudnZZBaIsUurnuoOjDEHqI3hVpijBO9ptv_FDku03udlncelu40Pln6ElruwTPH94TdPNu--XiQ3n18f3lxflV6SRTUwkW08pqIhtiG1BckJZL7mqlGVS21kor0VjiMqcwa2rOWUVrrp3m1jVA2Qk6XX3HGL7OkCbT--Sg6-wAYU6GCME1pyp_yP9RTpiSuiIZff0HehfmOOQgC4UZkZXGv6id7cD4oQ1TtG4xNedMaZGtBMvU2V-ofBrovQsDtD7PDwRvDgSZmeB-2tk5JXN5_fmQJSvrYkgpQmvG6Hsb94Zgs7TDrO0wuR1maYfZZ83Lh3Bz3UPzU_GjDhmgK5DyathB_C39P1xfraLWBmN30Sdzc00zkPumMqvYd00By9A</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>1540316890</pqid></control><display><type>article</type><title>Expression and purification of soluble monomeric streptavidin in Escherichia coli</title><source>MEDLINE</source><source>SpringerLink Journals (MCLS)</source><creator>Demonte, Daniel ; Dundas, Christopher M ; Park, Sheldon</creator><creatorcontrib>Demonte, Daniel ; Dundas, Christopher M ; Park, Sheldon</creatorcontrib><description>We recently reported the engineering of monomeric streptavidin (mSA) for use in monomeric detection of biotinylated ligands. Although mSA can be expressed functionally on the surface of mammalian cells and yeast, the molecule does not fold correctly when expressed in Escherichia coli. Refolding from inclusion bodies is cumbersome and yields a limited amount of purified protein. Improving the final yield should facilitate its use in biotechnology. We tested the expression and purification of mSA fused to GST, MBP, thioredoxin, and sumo tags to simplify its purification and improve the yield. The fusion proteins can be expressed solubly in E. coli and increase the yield by more than 20-fold. Unmodified mSA can be obtained by proteolytically removing the fusion tag. Purified mSA can be immobilized on a solid matrix to purify biotinylated ligands. Together, expressing mSA as a fusion with a solubilization tag vastly simplifies its preparation and increases its usability in biotechnology.</description><identifier>ISSN: 0175-7598</identifier><identifier>EISSN: 1432-0614</identifier><identifier>DOI: 10.1007/s00253-014-5682-y</identifier><identifier>PMID: 24691867</identifier><language>eng</language><publisher>Berlin/Heidelberg: Springer-Verlag</publisher><subject>Analysis ; Biomedical and Life Sciences ; Biosynthesis ; Biotechnological Products and Process Engineering ; Biotechnology ; Chemical engineering ; Cloning ; E coli ; engineering ; Escherichia coli ; Escherichia coli - genetics ; Escherichia coli - metabolism ; Gene Expression ; inclusion bodies ; Labeling ; Life Sciences ; Ligands ; mammals ; Methods ; Microbial Genetics and Genomics ; Microbiological synthesis ; Microbiology ; Microorganisms ; Molecular probes ; Physiological aspects ; Proteins ; Recombinant Fusion Proteins - biosynthesis ; Recombinant Fusion Proteins - chemistry ; Recombinant Fusion Proteins - genetics ; Solubility ; solubilization ; streptavidin ; Streptavidin - biosynthesis ; Streptavidin - chemistry ; Streptavidin - genetics ; Studies ; Yeasts</subject><ispartof>Applied microbiology and biotechnology, 2014, Vol.98 (14), p.6285-6295</ispartof><rights>Springer-Verlag Berlin Heidelberg 2014</rights><rights>COPYRIGHT 2014 Springer</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c637t-ea028a916d1ade7451f464cb793e8ab97975da1cea0703db44382b49c94acde23</citedby><cites>FETCH-LOGICAL-c637t-ea028a916d1ade7451f464cb793e8ab97975da1cea0703db44382b49c94acde23</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1007/s00253-014-5682-y$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s00253-014-5682-y$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,776,780,27901,27902,41464,42533,51294</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24691867$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Demonte, Daniel</creatorcontrib><creatorcontrib>Dundas, Christopher M</creatorcontrib><creatorcontrib>Park, Sheldon</creatorcontrib><title>Expression and purification of soluble monomeric streptavidin in Escherichia coli</title><title>Applied microbiology and biotechnology</title><addtitle>Appl Microbiol Biotechnol</addtitle><addtitle>Appl Microbiol Biotechnol</addtitle><description>We recently reported the engineering of monomeric streptavidin (mSA) for use in monomeric detection of biotinylated ligands. Although mSA can be expressed functionally on the surface of mammalian cells and yeast, the molecule does not fold correctly when expressed in Escherichia coli. Refolding from inclusion bodies is cumbersome and yields a limited amount of purified protein. Improving the final yield should facilitate its use in biotechnology. We tested the expression and purification of mSA fused to GST, MBP, thioredoxin, and sumo tags to simplify its purification and improve the yield. The fusion proteins can be expressed solubly in E. coli and increase the yield by more than 20-fold. Unmodified mSA can be obtained by proteolytically removing the fusion tag. Purified mSA can be immobilized on a solid matrix to purify biotinylated ligands. Together, expressing mSA as a fusion with a solubilization tag vastly simplifies its preparation and increases its usability in biotechnology.</description><subject>Analysis</subject><subject>Biomedical and Life Sciences</subject><subject>Biosynthesis</subject><subject>Biotechnological Products and Process Engineering</subject><subject>Biotechnology</subject><subject>Chemical engineering</subject><subject>Cloning</subject><subject>E coli</subject><subject>engineering</subject><subject>Escherichia coli</subject><subject>Escherichia coli - genetics</subject><subject>Escherichia coli - metabolism</subject><subject>Gene Expression</subject><subject>inclusion bodies</subject><subject>Labeling</subject><subject>Life Sciences</subject><subject>Ligands</subject><subject>mammals</subject><subject>Methods</subject><subject>Microbial Genetics and Genomics</subject><subject>Microbiological synthesis</subject><subject>Microbiology</subject><subject>Microorganisms</subject><subject>Molecular probes</subject><subject>Physiological aspects</subject><subject>Proteins</subject><subject>Recombinant Fusion Proteins - biosynthesis</subject><subject>Recombinant Fusion Proteins - chemistry</subject><subject>Recombinant Fusion Proteins - genetics</subject><subject>Solubility</subject><subject>solubilization</subject><subject>streptavidin</subject><subject>Streptavidin - biosynthesis</subject><subject>Streptavidin - chemistry</subject><subject>Streptavidin - genetics</subject><subject>Studies</subject><subject>Yeasts</subject><issn>0175-7598</issn><issn>1432-0614</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2014</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>BENPR</sourceid><recordid>eNqFkt1r1TAYxoMo7mz6B3ijBW_mRWe-01yOcdTBQHTuOqTp27OMtqlJKzv_vSmdH0dESSDwvr_ngSc8CL0g-IxgrN4mjKlgJSa8FLKi5f4R2hDOaIkl4Y_RBhMlSiV0dYSOU7rDmNBKyqfoiHKpSSXVBn3a3o8RUvJhKOzQFOMcfeudnZZBaIsUurnuoOjDEHqI3hVpijBO9ptv_FDku03udlncelu40Pln6ElruwTPH94TdPNu--XiQ3n18f3lxflV6SRTUwkW08pqIhtiG1BckJZL7mqlGVS21kor0VjiMqcwa2rOWUVrrp3m1jVA2Qk6XX3HGL7OkCbT--Sg6-wAYU6GCME1pyp_yP9RTpiSuiIZff0HehfmOOQgC4UZkZXGv6id7cD4oQ1TtG4xNedMaZGtBMvU2V-ofBrovQsDtD7PDwRvDgSZmeB-2tk5JXN5_fmQJSvrYkgpQmvG6Hsb94Zgs7TDrO0wuR1maYfZZ83Lh3Bz3UPzU_GjDhmgK5DyathB_C39P1xfraLWBmN30Sdzc00zkPumMqvYd00By9A</recordid><startdate>2014</startdate><enddate>2014</enddate><creator>Demonte, Daniel</creator><creator>Dundas, Christopher M</creator><creator>Park, Sheldon</creator><general>Springer-Verlag</general><general>Springer Berlin Heidelberg</general><general>Springer</general><general>Springer Nature B.V</general><scope>FBQ</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>ISR</scope><scope>3V.</scope><scope>7QL</scope><scope>7T7</scope><scope>7WY</scope><scope>7WZ</scope><scope>7X7</scope><scope>7XB</scope><scope>87Z</scope><scope>88A</scope><scope>88E</scope><scope>88I</scope><scope>8AO</scope><scope>8FD</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>8FL</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BEZIV</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FRNLG</scope><scope>FYUFA</scope><scope>F~G</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K60</scope><scope>K6~</scope><scope>K9.</scope><scope>L.-</scope><scope>LK8</scope><scope>M0C</scope><scope>M0S</scope><scope>M1P</scope><scope>M2P</scope><scope>M7N</scope><scope>M7P</scope><scope>P64</scope><scope>PQBIZ</scope><scope>PQBZA</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>Q9U</scope><scope>7X8</scope><scope>7QO</scope></search><sort><creationdate>2014</creationdate><title>Expression and purification of soluble monomeric streptavidin in Escherichia coli</title><author>Demonte, Daniel ; Dundas, Christopher M ; Park, Sheldon</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c637t-ea028a916d1ade7451f464cb793e8ab97975da1cea0703db44382b49c94acde23</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2014</creationdate><topic>Analysis</topic><topic>Biomedical and Life Sciences</topic><topic>Biosynthesis</topic><topic>Biotechnological Products and Process Engineering</topic><topic>Biotechnology</topic><topic>Chemical engineering</topic><topic>Cloning</topic><topic>E coli</topic><topic>engineering</topic><topic>Escherichia coli</topic><topic>Escherichia coli - genetics</topic><topic>Escherichia coli - metabolism</topic><topic>Gene Expression</topic><topic>inclusion bodies</topic><topic>Labeling</topic><topic>Life Sciences</topic><topic>Ligands</topic><topic>mammals</topic><topic>Methods</topic><topic>Microbial Genetics and Genomics</topic><topic>Microbiological synthesis</topic><topic>Microbiology</topic><topic>Microorganisms</topic><topic>Molecular probes</topic><topic>Physiological aspects</topic><topic>Proteins</topic><topic>Recombinant Fusion Proteins - biosynthesis</topic><topic>Recombinant Fusion Proteins - chemistry</topic><topic>Recombinant Fusion Proteins - genetics</topic><topic>Solubility</topic><topic>solubilization</topic><topic>streptavidin</topic><topic>Streptavidin - biosynthesis</topic><topic>Streptavidin - chemistry</topic><topic>Streptavidin - genetics</topic><topic>Studies</topic><topic>Yeasts</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Demonte, Daniel</creatorcontrib><creatorcontrib>Dundas, Christopher M</creatorcontrib><creatorcontrib>Park, Sheldon</creatorcontrib><collection>AGRIS</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Gale In Context: Science</collection><collection>ProQuest Central (Corporate)</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>ABI/INFORM Collection</collection><collection>ABI/INFORM Global (PDF only)</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>ABI/INFORM Global (Alumni Edition)</collection><collection>Biology Database (Alumni Edition)</collection><collection>Medical Database (Alumni Edition)</collection><collection>Science Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>Technology Research Database</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ABI/INFORM Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest One Sustainability</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Business Premium Collection</collection><collection>Natural Science Collection</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>Engineering Research Database</collection><collection>Business Premium Collection (Alumni)</collection><collection>Health Research Premium Collection</collection><collection>ABI/INFORM Global (Corporate)</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Business Collection (Alumni Edition)</collection><collection>ProQuest Business Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>ABI/INFORM Professional Advanced</collection><collection>ProQuest Biological Science Collection</collection><collection>ABI/INFORM Global</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Science Database</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biological Science Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>One Business (ProQuest)</collection><collection>ProQuest One Business (Alumni)</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central Basic</collection><collection>MEDLINE - Academic</collection><collection>Biotechnology Research Abstracts</collection><jtitle>Applied microbiology and biotechnology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Demonte, Daniel</au><au>Dundas, Christopher M</au><au>Park, Sheldon</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Expression and purification of soluble monomeric streptavidin in Escherichia coli</atitle><jtitle>Applied microbiology and biotechnology</jtitle><stitle>Appl Microbiol Biotechnol</stitle><addtitle>Appl Microbiol Biotechnol</addtitle><date>2014</date><risdate>2014</risdate><volume>98</volume><issue>14</issue><spage>6285</spage><epage>6295</epage><pages>6285-6295</pages><issn>0175-7598</issn><eissn>1432-0614</eissn><abstract>We recently reported the engineering of monomeric streptavidin (mSA) for use in monomeric detection of biotinylated ligands. Although mSA can be expressed functionally on the surface of mammalian cells and yeast, the molecule does not fold correctly when expressed in Escherichia coli. Refolding from inclusion bodies is cumbersome and yields a limited amount of purified protein. Improving the final yield should facilitate its use in biotechnology. We tested the expression and purification of mSA fused to GST, MBP, thioredoxin, and sumo tags to simplify its purification and improve the yield. The fusion proteins can be expressed solubly in E. coli and increase the yield by more than 20-fold. Unmodified mSA can be obtained by proteolytically removing the fusion tag. Purified mSA can be immobilized on a solid matrix to purify biotinylated ligands. Together, expressing mSA as a fusion with a solubilization tag vastly simplifies its preparation and increases its usability in biotechnology.</abstract><cop>Berlin/Heidelberg</cop><pub>Springer-Verlag</pub><pmid>24691867</pmid><doi>10.1007/s00253-014-5682-y</doi><tpages>11</tpages></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0175-7598 |
| ispartof | Applied microbiology and biotechnology, 2014, Vol.98 (14), p.6285-6295 |
| issn | 0175-7598 1432-0614 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_1554942714 |
| source | MEDLINE; SpringerLink Journals (MCLS) |
| subjects | Analysis Biomedical and Life Sciences Biosynthesis Biotechnological Products and Process Engineering Biotechnology Chemical engineering Cloning E coli engineering Escherichia coli Escherichia coli - genetics Escherichia coli - metabolism Gene Expression inclusion bodies Labeling Life Sciences Ligands mammals Methods Microbial Genetics and Genomics Microbiological synthesis Microbiology Microorganisms Molecular probes Physiological aspects Proteins Recombinant Fusion Proteins - biosynthesis Recombinant Fusion Proteins - chemistry Recombinant Fusion Proteins - genetics Solubility solubilization streptavidin Streptavidin - biosynthesis Streptavidin - chemistry Streptavidin - genetics Studies Yeasts |
| title | Expression and purification of soluble monomeric streptavidin in Escherichia coli |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-29T21%3A20%3A00IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-gale_proqu&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Expression%20and%20purification%20of%20soluble%20monomeric%20streptavidin%20in%20Escherichia%20coli&rft.jtitle=Applied%20microbiology%20and%20biotechnology&rft.au=Demonte,%20Daniel&rft.date=2014&rft.volume=98&rft.issue=14&rft.spage=6285&rft.epage=6295&rft.pages=6285-6295&rft.issn=0175-7598&rft.eissn=1432-0614&rft_id=info:doi/10.1007/s00253-014-5682-y&rft_dat=%3Cgale_proqu%3EA379569853%3C/gale_proqu%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=1540316890&rft_id=info:pmid/24691867&rft_galeid=A379569853&rfr_iscdi=true |