Protocol for aerosol-free recombinant production and NMR analysis of prion proteins
The central hallmark of prion diseases is the misfolding of cellular prion protein (PrP C ) into a disease-associated aggregated isoform known as scrapie prion protein (PrP Sc ). NMR spectroscopy has made many essential contributions to the characterization of recombinant PrP in its folded, unfolded...
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Veröffentlicht in: | Journal of biomolecular NMR 2014-06, Vol.59 (2), p.111-117 |
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creator | Rehbein, Peter Saxena, Krishna Schlepckow, Kai Schwalbe, Harald |
description | The central hallmark of prion diseases is the misfolding of cellular prion protein (PrP
C
) into a disease-associated aggregated isoform known as scrapie prion protein (PrP
Sc
). NMR spectroscopy has made many essential contributions to the characterization of recombinant PrP in its folded, unfolded and aggregated states. Recent studies reporting on de novo generation of prions from recombinant PrP and infection of animals using prion aerosols suggest that adjustment of current biosafety measures may be necessary, particularly given the relatively high protein concentrations required for NMR applications that favor aggregation. We here present a protocol for the production of recombinant PrP under biosafety level 2 conditions that avoids entirely exposure of the experimenter to aerosols that might contain harmful PrP aggregates. In addition, we introduce an NMR sample tube setup that allows for safe handling of PrP samples at the spectrometer that usually is not part of a dedicated biosafety level 2 laboratory. |
doi_str_mv | 10.1007/s10858-014-9831-5 |
format | Article |
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C
) into a disease-associated aggregated isoform known as scrapie prion protein (PrP
Sc
). NMR spectroscopy has made many essential contributions to the characterization of recombinant PrP in its folded, unfolded and aggregated states. Recent studies reporting on de novo generation of prions from recombinant PrP and infection of animals using prion aerosols suggest that adjustment of current biosafety measures may be necessary, particularly given the relatively high protein concentrations required for NMR applications that favor aggregation. We here present a protocol for the production of recombinant PrP under biosafety level 2 conditions that avoids entirely exposure of the experimenter to aerosols that might contain harmful PrP aggregates. In addition, we introduce an NMR sample tube setup that allows for safe handling of PrP samples at the spectrometer that usually is not part of a dedicated biosafety level 2 laboratory.</description><identifier>ISSN: 0925-2738</identifier><identifier>EISSN: 1573-5001</identifier><identifier>DOI: 10.1007/s10858-014-9831-5</identifier><identifier>PMID: 24771297</identifier><language>eng</language><publisher>Dordrecht: Springer Netherlands</publisher><subject>Aerosols ; Algorithms ; Amino Acid Sequence ; Amino Acids - chemistry ; Biochemistry ; Biological and Medical Physics ; Biophysics ; Biosafety ; Computer Simulation ; Molecular Sequence Data ; Nuclear Magnetic Resonance, Biomolecular ; Physics ; Physics and Astronomy ; Prions - chemistry ; Recombinant Proteins - chemistry ; Recombinant Proteins - metabolism ; Spectroscopy/Spectrometry</subject><ispartof>Journal of biomolecular NMR, 2014-06, Vol.59 (2), p.111-117</ispartof><rights>Springer Science+Business Media Dordrecht 2014</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c405t-40ea7f23951348195fb75c5ebc115c725a2493b734b78d48e866523fef4c22a43</citedby><cites>FETCH-LOGICAL-c405t-40ea7f23951348195fb75c5ebc115c725a2493b734b78d48e866523fef4c22a43</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/s10858-014-9831-5$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s10858-014-9831-5$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,780,784,27924,27925,41488,42557,51319</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24771297$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Rehbein, Peter</creatorcontrib><creatorcontrib>Saxena, Krishna</creatorcontrib><creatorcontrib>Schlepckow, Kai</creatorcontrib><creatorcontrib>Schwalbe, Harald</creatorcontrib><title>Protocol for aerosol-free recombinant production and NMR analysis of prion proteins</title><title>Journal of biomolecular NMR</title><addtitle>J Biomol NMR</addtitle><addtitle>J Biomol NMR</addtitle><description>The central hallmark of prion diseases is the misfolding of cellular prion protein (PrP
C
) into a disease-associated aggregated isoform known as scrapie prion protein (PrP
Sc
). NMR spectroscopy has made many essential contributions to the characterization of recombinant PrP in its folded, unfolded and aggregated states. Recent studies reporting on de novo generation of prions from recombinant PrP and infection of animals using prion aerosols suggest that adjustment of current biosafety measures may be necessary, particularly given the relatively high protein concentrations required for NMR applications that favor aggregation. We here present a protocol for the production of recombinant PrP under biosafety level 2 conditions that avoids entirely exposure of the experimenter to aerosols that might contain harmful PrP aggregates. In addition, we introduce an NMR sample tube setup that allows for safe handling of PrP samples at the spectrometer that usually is not part of a dedicated biosafety level 2 laboratory.</description><subject>Aerosols</subject><subject>Algorithms</subject><subject>Amino Acid Sequence</subject><subject>Amino Acids - chemistry</subject><subject>Biochemistry</subject><subject>Biological and Medical Physics</subject><subject>Biophysics</subject><subject>Biosafety</subject><subject>Computer Simulation</subject><subject>Molecular Sequence Data</subject><subject>Nuclear Magnetic Resonance, Biomolecular</subject><subject>Physics</subject><subject>Physics and Astronomy</subject><subject>Prions - chemistry</subject><subject>Recombinant Proteins - chemistry</subject><subject>Recombinant Proteins - metabolism</subject><subject>Spectroscopy/Spectrometry</subject><issn>0925-2738</issn><issn>1573-5001</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2014</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNqNkc1PGzEQxS1ERULaP4BLtRKXXtyOPyb2HlEEBSkU1I-z5XVstNFmXezdQ_77OiSgCgmJ0xzeb95o3iPkjMFXBqC-ZQYaNQUmaa0Fo3hEpgyVoAjAjskUao6UK6En5DTnNQDUms9PyIRLpRiv1ZT8uk9xiC52VYipsj7FHDsakvdV8i5umra3_VD9TXE1uqGNfWX7VfXj9meZttvmNlcxFHmnFGjwbZ8_kg_Bdtl_OswZ-XN1-XtxTZd3328WF0vqJOBAJXirAhc1MiE1qzE0Ch36xjGGTnG0XNaiUUI2Sq-k9no-Ry6CD9JxbqWYkS9733L4cfR5MJs2O991tvdxzIahlCUmEPAOlM-VQARW0PNX6DqOqTz7RKGui-nuNttTriSWkw-mZLCxaWsYmF05Zl-OKeWYXTkGy87ng_PYbPzqZeO5jQLwPZCL1D_49N_pN13_AWE3mBc</recordid><startdate>20140601</startdate><enddate>20140601</enddate><creator>Rehbein, Peter</creator><creator>Saxena, Krishna</creator><creator>Schlepckow, Kai</creator><creator>Schwalbe, Harald</creator><general>Springer Netherlands</general><general>Springer Nature B.V</general><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>3V.</scope><scope>7QL</scope><scope>7QO</scope><scope>7TK</scope><scope>7TM</scope><scope>7U9</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>ARAPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M7P</scope><scope>P5Z</scope><scope>P62</scope><scope>P64</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>7X8</scope></search><sort><creationdate>20140601</creationdate><title>Protocol for aerosol-free recombinant production and NMR analysis of prion proteins</title><author>Rehbein, Peter ; 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C
) into a disease-associated aggregated isoform known as scrapie prion protein (PrP
Sc
). NMR spectroscopy has made many essential contributions to the characterization of recombinant PrP in its folded, unfolded and aggregated states. Recent studies reporting on de novo generation of prions from recombinant PrP and infection of animals using prion aerosols suggest that adjustment of current biosafety measures may be necessary, particularly given the relatively high protein concentrations required for NMR applications that favor aggregation. We here present a protocol for the production of recombinant PrP under biosafety level 2 conditions that avoids entirely exposure of the experimenter to aerosols that might contain harmful PrP aggregates. In addition, we introduce an NMR sample tube setup that allows for safe handling of PrP samples at the spectrometer that usually is not part of a dedicated biosafety level 2 laboratory.</abstract><cop>Dordrecht</cop><pub>Springer Netherlands</pub><pmid>24771297</pmid><doi>10.1007/s10858-014-9831-5</doi><tpages>7</tpages></addata></record> |
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subjects | Aerosols Algorithms Amino Acid Sequence Amino Acids - chemistry Biochemistry Biological and Medical Physics Biophysics Biosafety Computer Simulation Molecular Sequence Data Nuclear Magnetic Resonance, Biomolecular Physics Physics and Astronomy Prions - chemistry Recombinant Proteins - chemistry Recombinant Proteins - metabolism Spectroscopy/Spectrometry |
title | Protocol for aerosol-free recombinant production and NMR analysis of prion proteins |
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