Hin recombinase assembles a tetrameric protein swivel that exchanges DNA strands
Most site-specific recombinases can be grouped into two structurally and mechanistically different classes. Whereas recombination by tyrosine recombinases proceeds with little movements by the proteins, serine recombinases exchange DNA strands by a mechanism requiring large quaternary rearrangements...
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Veröffentlicht in: | Nucleic acids research 2009-08, Vol.37 (14), p.4743-4756 |
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creator | Dhar, Gautam McLean, Meghan M Heiss, John K Johnson, Reid C |
description | Most site-specific recombinases can be grouped into two structurally and mechanistically different classes. Whereas recombination by tyrosine recombinases proceeds with little movements by the proteins, serine recombinases exchange DNA strands by a mechanism requiring large quaternary rearrangements. Here we use site-directed crosslinking to investigate the conformational changes that accompany the formation of the synaptic complex and the exchange of DNA strands by the Hin serine recombinase. Efficient crosslinking between residues corresponding to the 'D-helix' region provides the first experimental evidence for interactions between synapsed subunits within this region and distinguishes between different tetrameric conformers that have been observed in crystal structures of related serine recombinases. Crosslinking profiles between cysteines introduced over the 35 residue E-helix region that constitutes most of the proposed rotating interface both support the long helical structure of the region and provide strong experimental support for a subunit rotation mechanism that mediates DNA exchange. |
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Whereas recombination by tyrosine recombinases proceeds with little movements by the proteins, serine recombinases exchange DNA strands by a mechanism requiring large quaternary rearrangements. Here we use site-directed crosslinking to investigate the conformational changes that accompany the formation of the synaptic complex and the exchange of DNA strands by the Hin serine recombinase. Efficient crosslinking between residues corresponding to the 'D-helix' region provides the first experimental evidence for interactions between synapsed subunits within this region and distinguishes between different tetrameric conformers that have been observed in crystal structures of related serine recombinases. 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Whereas recombination by tyrosine recombinases proceeds with little movements by the proteins, serine recombinases exchange DNA strands by a mechanism requiring large quaternary rearrangements. Here we use site-directed crosslinking to investigate the conformational changes that accompany the formation of the synaptic complex and the exchange of DNA strands by the Hin serine recombinase. Efficient crosslinking between residues corresponding to the 'D-helix' region provides the first experimental evidence for interactions between synapsed subunits within this region and distinguishes between different tetrameric conformers that have been observed in crystal structures of related serine recombinases. Crosslinking profiles between cysteines introduced over the 35 residue E-helix region that constitutes most of the proposed rotating interface both support the long helical structure of the region and provide strong experimental support for a subunit rotation mechanism that mediates DNA exchange.</description><subject>Amino Acid Sequence</subject><subject>Cross-Linking Reagents</subject><subject>Cysteine - chemistry</subject><subject>DNA - chemistry</subject><subject>DNA - metabolism</subject><subject>DNA Nucleotidyltransferases - chemistry</subject><subject>DNA Nucleotidyltransferases - genetics</subject><subject>DNA Nucleotidyltransferases - metabolism</subject><subject>Models, Molecular</subject><subject>Molecular Sequence Data</subject><subject>Mutation</subject><subject>Nucleic Acid Enzymes</subject><subject>Protein Multimerization</subject><subject>Protein Subunits - chemistry</subject><subject>Protein Subunits - metabolism</subject><subject>Recombinases - chemistry</subject><subject>Recombinases - genetics</subject><subject>Recombinases - metabolism</subject><issn>0305-1048</issn><issn>1362-4962</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</creationdate><recordtype>article</recordtype><sourceid>TOX</sourceid><sourceid>EIF</sourceid><recordid>eNqF0ctu1DAUBmALUdGhsOEBIEKCRaVQ3xNvKpXSMsCoIEEl1I1lJyczbpM42Elp3x6jjMplASsv_Om3z_kRekLwK4IVO-hNOFhfDVzKe2hBmKQ5V5LeRwvMsMgJ5uUuehjjJcaEE8EfoF2iBBGKsQX6tHR9FqDynXW9iZCZGKGzLcTMZCOMwXQQXJUNwY-QaPzurqHNxo0ZM7ipNqZfJ_rm7CiLyfZ1fIR2GtNGeLw999D56cmX42W--vj23fHRKq8ElWNuGS1FWTeWiFo0jZS8LKU1mFjWNBi4AloyWwGTxNKSgFTMcmOsKiiugVu2hw7n3GGyHdQV9On9Vg_BdSbcam-c_vOmdxu99teaFpTTkqaAl9uA4L9NEEfduVhB25oe_BS1LETBSSH-CykuFMacJfj8L3jpp9CnLSSDJWWCqoT2Z1QFH2OA5u7LBOufdepUp57rTPjp70P-otv-EngxAz8N_w7KZ-fiCDd30oSrNCcrhF5-vdAfyPvX-Oz0Qq-Sfzb7xnht1sFFff6ZYsIwkVIWJWc_AMkFwgI</recordid><startdate>20090801</startdate><enddate>20090801</enddate><creator>Dhar, Gautam</creator><creator>McLean, Meghan M</creator><creator>Heiss, John K</creator><creator>Johnson, Reid C</creator><general>Oxford University Press</general><general>Oxford Publishing Limited (England)</general><scope>FBQ</scope><scope>BSCLL</scope><scope>TOX</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>7QP</scope><scope>7QR</scope><scope>7SS</scope><scope>7TK</scope><scope>7TM</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>K9.</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20090801</creationdate><title>Hin recombinase assembles a tetrameric protein swivel that exchanges DNA strands</title><author>Dhar, Gautam ; 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Whereas recombination by tyrosine recombinases proceeds with little movements by the proteins, serine recombinases exchange DNA strands by a mechanism requiring large quaternary rearrangements. Here we use site-directed crosslinking to investigate the conformational changes that accompany the formation of the synaptic complex and the exchange of DNA strands by the Hin serine recombinase. Efficient crosslinking between residues corresponding to the 'D-helix' region provides the first experimental evidence for interactions between synapsed subunits within this region and distinguishes between different tetrameric conformers that have been observed in crystal structures of related serine recombinases. 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subjects | Amino Acid Sequence Cross-Linking Reagents Cysteine - chemistry DNA - chemistry DNA - metabolism DNA Nucleotidyltransferases - chemistry DNA Nucleotidyltransferases - genetics DNA Nucleotidyltransferases - metabolism Models, Molecular Molecular Sequence Data Mutation Nucleic Acid Enzymes Protein Multimerization Protein Subunits - chemistry Protein Subunits - metabolism Recombinases - chemistry Recombinases - genetics Recombinases - metabolism |
title | Hin recombinase assembles a tetrameric protein swivel that exchanges DNA strands |
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