The 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. 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.
Dhar
Gautam
G
Department of Biological Chemistry, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, CA 90095, USA.
McLean
Meghan M
MM
Heiss
John K
JK
Johnson
Reid C
RC
D000595:Amino Acid Sequence; D003432:Cross-Linking Reagents; D003545:Cysteine / Q000737:chemistry; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004254:DNA Nucleotidyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D045522:Recombinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D003432:Cross-Linking Reagents; D021122:Protein Subunits; D045522:Recombinases; D004247:DNA; D004254:DNA Nucleotidyltransferases; C059997:Hin recombinase; D003545:Cysteine