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spinqualitylabelsall models 1nzb, resolution 3.10Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

Crystal structure of wild type Cre recombinase-loxP synapse

Overview

Escherichia coli phage P1 Cre recombinase catalyzes the site-specific, recombination of DNA containing loxP sites. We report here two crystal, structures of a wild-type Cre recombinase-loxP synaptic complex, corresponding to two distinct reaction states: an initial pre-cleavage, complex, trapped using a phosphorothioate modification at the cleavable, scissile bond that prevents the recombination reaction, and a, 3'-phosphotyrosine protein-DNA intermediate resulting from the first, strand cleavage. In contrast to previously determined Cre complexes, both, structures contain a full tetrameric complex in the asymmetric unit, unequivocally showing that the anti-parallel arrangement of the loxP sites, is an intrinsic property of the Cre-loxP recombination synapse. The, conformation of the spacer is different to the one observed for the, symmetrized loxS site: a kink next to the scissile phosphate in the top, strand of the pre-cleavage complex leads to unstacking of the TpG step and, a widening of the minor groove. This side of the spacer is interacting, with a 'cleavage-competent' Cre subunit, suggesting that the first, cleavage occurs at the ApT step in the top strand. This is further, confirmed by the structure of the 3'-phosphotyrosine intermediate, where, the DNA is cleaved in the top strands and covalently linked to the, 'cleavage-competent' subunits. The cleavage is followed by a movement of, the C-terminal part containing the attacking Y324 and the helix N, interacting with the 'non-cleaving' subunit. This rearrangement could be, responsible for the interconversion of Cre subunits. Our results also, suggest that the Cre-induced kink next to the scissile phosphodiester, activates the DNA for cleavage at this position and facilitates strand, transfer.

About this Structure

1NZB is a Single protein structure of sequence from Enterobacteria phage p21 with IOD and MG as ligands. Full crystallographic information is available from OCA.

Reference

Crystal structure of a wild-type Cre recombinase-loxP synapse reveals a novel spacer conformation suggesting an alternative mechanism for DNA cleavage activation., Ennifar E, Meyer JE, Buchholz F, Stewart AF, Suck D, Nucleic Acids Res. 2003 Sep 15;31(18):5449-60. PMID:12954782

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