Chromophore-assisted light inactivation (CALI) using the phototoxic fluorescent protein KillerRed
The phototoxic red fluorescent GFP-like protein KillerRed has recently been described. The phototoxicity of KillerRed exceeds that of EGFP by at least 1,000-fold, making it the first fully genetically encoded photosensitizer. KillerRed opens up new possibilities for precise light-induced cell killin...
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| Veröffentlicht in: | Nature protocols 2006-08, Vol.1 (2), p.947-953 |
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| creator | Lukyanov, Konstantin A Bulina, Maria E Britanova, Olga V Onichtchouk, Daria Lukyanov, Sergey Chudakov, Dmitriy M |
| description | The phototoxic red fluorescent GFP-like protein KillerRed has recently been described. The phototoxicity of KillerRed exceeds that of EGFP by at least 1,000-fold, making it the first fully genetically encoded photosensitizer. KillerRed opens up new possibilities for precise light-induced cell killing and target protein inactivation. Because KillerRed is encoded by a gene, it can be expressed in a spatially and temporally regulated manner, under a chosen promoter, and fused with the desired protein of interest or localization signal. Here we provide a protocol for target protein inactivation in cell culture using KillerRed. As KillerRed is a new tool, the protocol focuses on aspects that will allow users to maximize the potential of this protein, guiding the design of chimeric constructs, recommended control experiments and preferred illumination parameters. The protocol, which describes target protein visualization and subsequent inactivation, is a 2- or 3-d procedure. |
| doi_str_mv | 10.1038/nprot.2006.89 |
| format | Article |
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The phototoxicity of KillerRed exceeds that of EGFP by at least 1,000-fold, making it the first fully genetically encoded photosensitizer. KillerRed opens up new possibilities for precise light-induced cell killing and target protein inactivation. Because KillerRed is encoded by a gene, it can be expressed in a spatially and temporally regulated manner, under a chosen promoter, and fused with the desired protein of interest or localization signal. Here we provide a protocol for target protein inactivation in cell culture using KillerRed. As KillerRed is a new tool, the protocol focuses on aspects that will allow users to maximize the potential of this protein, guiding the design of chimeric constructs, recommended control experiments and preferred illumination parameters. The protocol, which describes target protein visualization and subsequent inactivation, is a 2- or 3-d procedure.</description><identifier>ISSN: 1754-2189</identifier><identifier>EISSN: 1750-2799</identifier><identifier>DOI: 10.1038/nprot.2006.89</identifier><identifier>PMID: 17406328</identifier><language>eng</language><publisher>London: Nature Publishing Group UK</publisher><subject>Active oxygen ; Analytical Chemistry ; Bacteria - genetics ; Bacteria - metabolism ; Biological Techniques ; Biomedical and Life Sciences ; Chromophores ; Computational Biology/Bioinformatics ; Diagnostic imaging ; Epithelial Cells - cytology ; Epithelial Cells - metabolism ; Gene Expression Regulation ; Green Fluorescent Proteins - chemistry ; Green Fluorescent Proteins - metabolism ; HeLa Cells ; Humans ; Inactivation ; Life Sciences ; Light ; Localization ; Luminescent Agents - chemistry ; Luminescent Agents - metabolism ; Methods ; Microarrays ; Organic Chemistry ; Phototoxicity ; Physiological aspects ; Protein Binding ; Proteins ; Protocol</subject><ispartof>Nature protocols, 2006-08, Vol.1 (2), p.947-953</ispartof><rights>Springer Nature Limited 2006</rights><rights>COPYRIGHT 2006 Nature Publishing Group</rights><rights>Copyright Nature Publishing Group Jun 2006</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c589t-ce659e1311e843ca1c6eae43ac7b21aaed8e39c73ad4928de0bdd7a1120b74f53</citedby><cites>FETCH-LOGICAL-c589t-ce659e1311e843ca1c6eae43ac7b21aaed8e39c73ad4928de0bdd7a1120b74f53</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,2727,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/17406328$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Lukyanov, Konstantin A</creatorcontrib><creatorcontrib>Bulina, Maria E</creatorcontrib><creatorcontrib>Britanova, Olga V</creatorcontrib><creatorcontrib>Onichtchouk, Daria</creatorcontrib><creatorcontrib>Lukyanov, Sergey</creatorcontrib><creatorcontrib>Chudakov, Dmitriy M</creatorcontrib><title>Chromophore-assisted light inactivation (CALI) using the phototoxic fluorescent protein KillerRed</title><title>Nature protocols</title><addtitle>Nat Protoc</addtitle><addtitle>Nat Protoc</addtitle><description>The phototoxic red fluorescent GFP-like protein KillerRed has recently been described. The phototoxicity of KillerRed exceeds that of EGFP by at least 1,000-fold, making it the first fully genetically encoded photosensitizer. KillerRed opens up new possibilities for precise light-induced cell killing and target protein inactivation. Because KillerRed is encoded by a gene, it can be expressed in a spatially and temporally regulated manner, under a chosen promoter, and fused with the desired protein of interest or localization signal. Here we provide a protocol for target protein inactivation in cell culture using KillerRed. As KillerRed is a new tool, the protocol focuses on aspects that will allow users to maximize the potential of this protein, guiding the design of chimeric constructs, recommended control experiments and preferred illumination parameters. The protocol, which describes target protein visualization and subsequent inactivation, is a 2- or 3-d procedure.</description><subject>Active oxygen</subject><subject>Analytical Chemistry</subject><subject>Bacteria - genetics</subject><subject>Bacteria - metabolism</subject><subject>Biological Techniques</subject><subject>Biomedical and Life Sciences</subject><subject>Chromophores</subject><subject>Computational Biology/Bioinformatics</subject><subject>Diagnostic imaging</subject><subject>Epithelial Cells - cytology</subject><subject>Epithelial Cells - metabolism</subject><subject>Gene Expression Regulation</subject><subject>Green Fluorescent Proteins - chemistry</subject><subject>Green Fluorescent Proteins - metabolism</subject><subject>HeLa Cells</subject><subject>Humans</subject><subject>Inactivation</subject><subject>Life Sciences</subject><subject>Light</subject><subject>Localization</subject><subject>Luminescent Agents - chemistry</subject><subject>Luminescent Agents - metabolism</subject><subject>Methods</subject><subject>Microarrays</subject><subject>Organic Chemistry</subject><subject>Phototoxicity</subject><subject>Physiological aspects</subject><subject>Protein Binding</subject><subject>Proteins</subject><subject>Protocol</subject><issn>1754-2189</issn><issn>1750-2799</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2006</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>eNp1kt1rFDEUxYMotlYffZUBQSwya752knlcFrVLi0Kt-BiymTuzKTPJmmSk_vdm26XrFMuFJCS_c7i5HIReEzwjmMmPbht8mlGMq5msn6BjIua4pKKun96eeUmJrI_QixivMeaCVeI5OiKC44pReYz0chP84LcbH6DUMdqYoCl6221SYZ02yf7WyXpXvF8uLlanxRit64q0gSJLUq4ba4q2H7M8GnCp2LUD1hXntu8hXELzEj1rdR_h1X4_QT8-f7panpUX376ssmlp5rJOpYFqXgNhhIDkzGhiKtDAmTZiTYnW0EhgtRFMN7ymsgG8bhqhCaF4LXg7Zyfo3Z1v7uDXCDGpweaW-l478GNUlWSU5yWDbx-A134MLvemCKOVYFISfqA63YOyrvUpaLOzVAsiKWaikjRTs_9QuRoYrPEOWpvvJ4LTiSAzCW5Sp8cY1er75ZT98Di7uPq5_DqlyzvaBB9jgFZtgx10-KMIVruoqNuoqF1UlKwz_2Y_hnE9QHOg99k4_C3mJ9dB-GdOjzjuR-Z0GgPcO06pv5Ws1LA</recordid><startdate>200608</startdate><enddate>200608</enddate><creator>Lukyanov, Konstantin A</creator><creator>Bulina, Maria E</creator><creator>Britanova, Olga V</creator><creator>Onichtchouk, Daria</creator><creator>Lukyanov, Sergey</creator><creator>Chudakov, Dmitriy M</creator><general>Nature Publishing Group UK</general><general>Nature Publishing Group</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>ATWCN</scope><scope>ISR</scope><scope>3V.</scope><scope>7QG</scope><scope>7T5</scope><scope>7T7</scope><scope>7TM</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FD</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</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>M7N</scope><scope>M7P</scope><scope>P64</scope><scope>PATMY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PYCSY</scope><scope>RC3</scope><scope>7X8</scope></search><sort><creationdate>200608</creationdate><title>Chromophore-assisted light inactivation (CALI) using the phototoxic fluorescent protein KillerRed</title><author>Lukyanov, Konstantin A ; Bulina, Maria E ; Britanova, Olga V ; Onichtchouk, Daria ; Lukyanov, Sergey ; Chudakov, Dmitriy M</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c589t-ce659e1311e843ca1c6eae43ac7b21aaed8e39c73ad4928de0bdd7a1120b74f53</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2006</creationdate><topic>Active oxygen</topic><topic>Analytical Chemistry</topic><topic>Bacteria - 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Academic</collection><jtitle>Nature protocols</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Lukyanov, Konstantin A</au><au>Bulina, Maria E</au><au>Britanova, Olga V</au><au>Onichtchouk, Daria</au><au>Lukyanov, Sergey</au><au>Chudakov, Dmitriy M</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Chromophore-assisted light inactivation (CALI) using the phototoxic fluorescent protein KillerRed</atitle><jtitle>Nature protocols</jtitle><stitle>Nat Protoc</stitle><addtitle>Nat Protoc</addtitle><date>2006-08</date><risdate>2006</risdate><volume>1</volume><issue>2</issue><spage>947</spage><epage>953</epage><pages>947-953</pages><issn>1754-2189</issn><eissn>1750-2799</eissn><abstract>The phototoxic red fluorescent GFP-like protein KillerRed has recently been described. The phototoxicity of KillerRed exceeds that of EGFP by at least 1,000-fold, making it the first fully genetically encoded photosensitizer. KillerRed opens up new possibilities for precise light-induced cell killing and target protein inactivation. Because KillerRed is encoded by a gene, it can be expressed in a spatially and temporally regulated manner, under a chosen promoter, and fused with the desired protein of interest or localization signal. Here we provide a protocol for target protein inactivation in cell culture using KillerRed. As KillerRed is a new tool, the protocol focuses on aspects that will allow users to maximize the potential of this protein, guiding the design of chimeric constructs, recommended control experiments and preferred illumination parameters. The protocol, which describes target protein visualization and subsequent inactivation, is a 2- or 3-d procedure.</abstract><cop>London</cop><pub>Nature Publishing Group UK</pub><pmid>17406328</pmid><doi>10.1038/nprot.2006.89</doi><tpages>7</tpages></addata></record> |
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| subjects | Active oxygen Analytical Chemistry Bacteria - genetics Bacteria - metabolism Biological Techniques Biomedical and Life Sciences Chromophores Computational Biology/Bioinformatics Diagnostic imaging Epithelial Cells - cytology Epithelial Cells - metabolism Gene Expression Regulation Green Fluorescent Proteins - chemistry Green Fluorescent Proteins - metabolism HeLa Cells Humans Inactivation Life Sciences Light Localization Luminescent Agents - chemistry Luminescent Agents - metabolism Methods Microarrays Organic Chemistry Phototoxicity Physiological aspects Protein Binding Proteins Protocol |
| title | Chromophore-assisted light inactivation (CALI) using the phototoxic fluorescent protein KillerRed |
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