Probing the Structure and Stability of a Hybrid Protein: The Human−E. coli Thioredoxin Chimera
The structure and stability of a hybrid protein composed of N-terminal human and C-terminal E. coli thioredoxin domains were investigated by NMR, fluorescence, and circular dichroism spectroscopy. We demonstrate that the chimeric protein is correctly folded and exhibits the common thioredoxin archit...
Gespeichert in:
| Veröffentlicht in: | Biochemistry (Easton) 2001-09, Vol.40 (37), p.11184-11192 |
|---|---|
| Hauptverfasser: | , , , , |
| Format: | Artikel |
| Sprache: | eng |
| Schlagworte: | |
| Online-Zugang: | Volltext |
| Tags: |
Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
|
| container_end_page | 11192 |
|---|---|
| container_issue | 37 |
| container_start_page | 11184 |
| container_title | Biochemistry (Easton) |
| container_volume | 40 |
| creator | Louis, John M Georgescu, Roxana E Tasayco, Maria Luisa Tcherkasskaya, Olga Gronenborn, Angela M |
| description | The structure and stability of a hybrid protein composed of N-terminal human and C-terminal E. coli thioredoxin domains were investigated by NMR, fluorescence, and circular dichroism spectroscopy. We demonstrate that the chimeric protein is correctly folded and exhibits the common thioredoxin architecture. However, the stability of the hybrid protein toward thermal and chemical denaturation is clearly reduced when compared with both parent proteins. Altogether, our data indicate that the interface between the two folding units of thioredoxin is tolerant toward changes in exact interdigitation of side chains, allowing for the formation of the unique overall thioredoxin fold. Further, the gene encoding the human−E. coli chimera was tested in vivo whether it supports the assembly of filamentous phages. No complementation of a thioredoxin-deficient E. coli mutant for the replication of the phages M13 or fd was observed, suggesting that parts of the overall protein structure in the N-terminal domain are crucial for this activity. |
| doi_str_mv | 10.1021/bi010745x |
| format | Article |
| fullrecord | <record><control><sourceid>proquest_cross</sourceid><recordid>TN_cdi_proquest_miscellaneous_71162963</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>71162963</sourcerecordid><originalsourceid>FETCH-LOGICAL-a349t-7f03ea3368018e384d74bb008365f760770ebf8f5776b60225b286d0c538b0f53</originalsourceid><addsrcrecordid>eNpt0EFP2zAUB3BrGloL22FfAPkyJA6B5zi2U26sAoJgWqV2u1p24lCXJGZ2IrU3jnDlI_JJ8NSKXXaynt_vvSf9EfpK4IRASk61BQIiY-sPaExYCkk2mbCPaAwAPEknHEZoP4RVLLPIPqERIYyRlIgxKmfeadvd4X5p8Lz3Q9kP3mDVVbFS2ja232BXY4WLjfa2wtH3xnZnr4_PeBFniqFV3evTy8UJLl1j45913lRubTs8XdrWePUZ7dWqCebL7j1Avy4vFtMiuf15dT09v00UzSZ9ImqgRlHKcyC5oXlWiUxrgJxyVgsOQoDRdV4zIbjmkKZMpzmvoGQ011AzeoCOtnsfvPszmNDL1obSNI3qjBuCFITwmAaN8HgLS-9C8KaWD962ym8kAfk3UfmeaLSHu6WDbk31T-4ijCDZAht6s37vK38vuaCCycVsLm--F79_zBZUFtF_23pVBrlyg-9iJv85_AZOwYwy</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>71162963</pqid></control><display><type>article</type><title>Probing the Structure and Stability of a Hybrid Protein: The Human−E. coli Thioredoxin Chimera</title><source>MEDLINE</source><source>American Chemical Society Journals</source><creator>Louis, John M ; Georgescu, Roxana E ; Tasayco, Maria Luisa ; Tcherkasskaya, Olga ; Gronenborn, Angela M</creator><creatorcontrib>Louis, John M ; Georgescu, Roxana E ; Tasayco, Maria Luisa ; Tcherkasskaya, Olga ; Gronenborn, Angela M</creatorcontrib><description>The structure and stability of a hybrid protein composed of N-terminal human and C-terminal E. coli thioredoxin domains were investigated by NMR, fluorescence, and circular dichroism spectroscopy. We demonstrate that the chimeric protein is correctly folded and exhibits the common thioredoxin architecture. However, the stability of the hybrid protein toward thermal and chemical denaturation is clearly reduced when compared with both parent proteins. Altogether, our data indicate that the interface between the two folding units of thioredoxin is tolerant toward changes in exact interdigitation of side chains, allowing for the formation of the unique overall thioredoxin fold. Further, the gene encoding the human−E. coli chimera was tested in vivo whether it supports the assembly of filamentous phages. No complementation of a thioredoxin-deficient E. coli mutant for the replication of the phages M13 or fd was observed, suggesting that parts of the overall protein structure in the N-terminal domain are crucial for this activity.</description><identifier>ISSN: 0006-2960</identifier><identifier>EISSN: 1520-4995</identifier><identifier>DOI: 10.1021/bi010745x</identifier><identifier>PMID: 11551217</identifier><language>eng</language><publisher>United States: American Chemical Society</publisher><subject>Amino Acid Sequence ; Bacterial Proteins - chemistry ; Escherichia coli ; Guanidine - pharmacology ; Hot Temperature ; Humans ; Models, Molecular ; Models, Theoretical ; Molecular Sequence Data ; Nuclear Magnetic Resonance, Biomolecular ; Protein Conformation ; Protein Denaturation ; Recombinant Fusion Proteins - chemistry ; Thermodynamics ; Thioredoxins - chemistry</subject><ispartof>Biochemistry (Easton), 2001-09, Vol.40 (37), p.11184-11192</ispartof><rights>Copyright © 2001 American Chemical Society</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a349t-7f03ea3368018e384d74bb008365f760770ebf8f5776b60225b286d0c538b0f53</citedby><cites>FETCH-LOGICAL-a349t-7f03ea3368018e384d74bb008365f760770ebf8f5776b60225b286d0c538b0f53</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://pubs.acs.org/doi/pdf/10.1021/bi010745x$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/bi010745x$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,780,784,2765,27076,27924,27925,56738,56788</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/11551217$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Louis, John M</creatorcontrib><creatorcontrib>Georgescu, Roxana E</creatorcontrib><creatorcontrib>Tasayco, Maria Luisa</creatorcontrib><creatorcontrib>Tcherkasskaya, Olga</creatorcontrib><creatorcontrib>Gronenborn, Angela M</creatorcontrib><title>Probing the Structure and Stability of a Hybrid Protein: The Human−E. coli Thioredoxin Chimera</title><title>Biochemistry (Easton)</title><addtitle>Biochemistry</addtitle><description>The structure and stability of a hybrid protein composed of N-terminal human and C-terminal E. coli thioredoxin domains were investigated by NMR, fluorescence, and circular dichroism spectroscopy. We demonstrate that the chimeric protein is correctly folded and exhibits the common thioredoxin architecture. However, the stability of the hybrid protein toward thermal and chemical denaturation is clearly reduced when compared with both parent proteins. Altogether, our data indicate that the interface between the two folding units of thioredoxin is tolerant toward changes in exact interdigitation of side chains, allowing for the formation of the unique overall thioredoxin fold. Further, the gene encoding the human−E. coli chimera was tested in vivo whether it supports the assembly of filamentous phages. No complementation of a thioredoxin-deficient E. coli mutant for the replication of the phages M13 or fd was observed, suggesting that parts of the overall protein structure in the N-terminal domain are crucial for this activity.</description><subject>Amino Acid Sequence</subject><subject>Bacterial Proteins - chemistry</subject><subject>Escherichia coli</subject><subject>Guanidine - pharmacology</subject><subject>Hot Temperature</subject><subject>Humans</subject><subject>Models, Molecular</subject><subject>Models, Theoretical</subject><subject>Molecular Sequence Data</subject><subject>Nuclear Magnetic Resonance, Biomolecular</subject><subject>Protein Conformation</subject><subject>Protein Denaturation</subject><subject>Recombinant Fusion Proteins - chemistry</subject><subject>Thermodynamics</subject><subject>Thioredoxins - chemistry</subject><issn>0006-2960</issn><issn>1520-4995</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2001</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpt0EFP2zAUB3BrGloL22FfAPkyJA6B5zi2U26sAoJgWqV2u1p24lCXJGZ2IrU3jnDlI_JJ8NSKXXaynt_vvSf9EfpK4IRASk61BQIiY-sPaExYCkk2mbCPaAwAPEknHEZoP4RVLLPIPqERIYyRlIgxKmfeadvd4X5p8Lz3Q9kP3mDVVbFS2ja232BXY4WLjfa2wtH3xnZnr4_PeBFniqFV3evTy8UJLl1j45913lRubTs8XdrWePUZ7dWqCebL7j1Avy4vFtMiuf15dT09v00UzSZ9ImqgRlHKcyC5oXlWiUxrgJxyVgsOQoDRdV4zIbjmkKZMpzmvoGQ011AzeoCOtnsfvPszmNDL1obSNI3qjBuCFITwmAaN8HgLS-9C8KaWD962ym8kAfk3UfmeaLSHu6WDbk31T-4ijCDZAht6s37vK38vuaCCycVsLm--F79_zBZUFtF_23pVBrlyg-9iJv85_AZOwYwy</recordid><startdate>20010918</startdate><enddate>20010918</enddate><creator>Louis, John M</creator><creator>Georgescu, Roxana E</creator><creator>Tasayco, Maria Luisa</creator><creator>Tcherkasskaya, Olga</creator><creator>Gronenborn, Angela M</creator><general>American Chemical Society</general><scope>BSCLL</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>7X8</scope></search><sort><creationdate>20010918</creationdate><title>Probing the Structure and Stability of a Hybrid Protein: The Human−E. coli Thioredoxin Chimera</title><author>Louis, John M ; Georgescu, Roxana E ; Tasayco, Maria Luisa ; Tcherkasskaya, Olga ; Gronenborn, Angela M</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a349t-7f03ea3368018e384d74bb008365f760770ebf8f5776b60225b286d0c538b0f53</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2001</creationdate><topic>Amino Acid Sequence</topic><topic>Bacterial Proteins - chemistry</topic><topic>Escherichia coli</topic><topic>Guanidine - pharmacology</topic><topic>Hot Temperature</topic><topic>Humans</topic><topic>Models, Molecular</topic><topic>Models, Theoretical</topic><topic>Molecular Sequence Data</topic><topic>Nuclear Magnetic Resonance, Biomolecular</topic><topic>Protein Conformation</topic><topic>Protein Denaturation</topic><topic>Recombinant Fusion Proteins - chemistry</topic><topic>Thermodynamics</topic><topic>Thioredoxins - chemistry</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Louis, John M</creatorcontrib><creatorcontrib>Georgescu, Roxana E</creatorcontrib><creatorcontrib>Tasayco, Maria Luisa</creatorcontrib><creatorcontrib>Tcherkasskaya, Olga</creatorcontrib><creatorcontrib>Gronenborn, Angela M</creatorcontrib><collection>Istex</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Biochemistry (Easton)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Louis, John M</au><au>Georgescu, Roxana E</au><au>Tasayco, Maria Luisa</au><au>Tcherkasskaya, Olga</au><au>Gronenborn, Angela M</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Probing the Structure and Stability of a Hybrid Protein: The Human−E. coli Thioredoxin Chimera</atitle><jtitle>Biochemistry (Easton)</jtitle><addtitle>Biochemistry</addtitle><date>2001-09-18</date><risdate>2001</risdate><volume>40</volume><issue>37</issue><spage>11184</spage><epage>11192</epage><pages>11184-11192</pages><issn>0006-2960</issn><eissn>1520-4995</eissn><abstract>The structure and stability of a hybrid protein composed of N-terminal human and C-terminal E. coli thioredoxin domains were investigated by NMR, fluorescence, and circular dichroism spectroscopy. We demonstrate that the chimeric protein is correctly folded and exhibits the common thioredoxin architecture. However, the stability of the hybrid protein toward thermal and chemical denaturation is clearly reduced when compared with both parent proteins. Altogether, our data indicate that the interface between the two folding units of thioredoxin is tolerant toward changes in exact interdigitation of side chains, allowing for the formation of the unique overall thioredoxin fold. Further, the gene encoding the human−E. coli chimera was tested in vivo whether it supports the assembly of filamentous phages. No complementation of a thioredoxin-deficient E. coli mutant for the replication of the phages M13 or fd was observed, suggesting that parts of the overall protein structure in the N-terminal domain are crucial for this activity.</abstract><cop>United States</cop><pub>American Chemical Society</pub><pmid>11551217</pmid><doi>10.1021/bi010745x</doi><tpages>9</tpages></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0006-2960 |
| ispartof | Biochemistry (Easton), 2001-09, Vol.40 (37), p.11184-11192 |
| issn | 0006-2960 1520-4995 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_71162963 |
| source | MEDLINE; American Chemical Society Journals |
| subjects | Amino Acid Sequence Bacterial Proteins - chemistry Escherichia coli Guanidine - pharmacology Hot Temperature Humans Models, Molecular Models, Theoretical Molecular Sequence Data Nuclear Magnetic Resonance, Biomolecular Protein Conformation Protein Denaturation Recombinant Fusion Proteins - chemistry Thermodynamics Thioredoxins - chemistry |
| title | Probing the Structure and Stability of a Hybrid Protein: The Human−E. coli Thioredoxin Chimera |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2024-12-25T02%3A51%3A37IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-proquest_cross&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Probing%20the%20Structure%20and%20Stability%20of%20a%20Hybrid%20Protein:%E2%80%89%20The%20Human%E2%88%92E.%20coli%20Thioredoxin%20Chimera&rft.jtitle=Biochemistry%20(Easton)&rft.au=Louis,%20John%20M&rft.date=2001-09-18&rft.volume=40&rft.issue=37&rft.spage=11184&rft.epage=11192&rft.pages=11184-11192&rft.issn=0006-2960&rft.eissn=1520-4995&rft_id=info:doi/10.1021/bi010745x&rft_dat=%3Cproquest_cross%3E71162963%3C/proquest_cross%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=71162963&rft_id=info:pmid/11551217&rfr_iscdi=true |