1KBU

CRE RECOMBINASE BOUND TO A LOXP HOLLIDAY JUNCTION

1KBU の概要

• エントリーDOI: 10.2210/pdb1kbu/pdb
• 関連するPDBエントリー: 1CRX 1DRG 1F44 2CRX 3CRX 4CRX
• 分子名称: LOXP, CRE RECOMBINASE, ... (4 entities in total)
• 機能のキーワード: site-specific recombinase, holliday junction complex, dna-protein co-crystal, int recombinase mechanism, hydrolase, ligase-dna complex, ligase/dna
• 由来する生物種: Enterobacteria phage P1
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 99758.66

構造登録者

Martin, S.S.,Pulido, E.,Chu, V.C.,Lechner, T.,Baldwin, E.P. (登録日: 2001-11-06, 公開日: 2002-06-07, 最終更新日: 2023-08-16)

主引用文献

Martin, S.S.,Pulido, E.,Chu, V.C.,Lechner, T.S.,Baldwin, E.P.
The Order of Strand Exchanges in Cre-LoxP Recombination and its Basis Suggested by the Crystal Structure of a Cre-LoxP Holliday Junction Complex
J.Mol.Biol., 319:107-127, 2002

PubMed Abstract: Cre recombinase uses two pairs of sequential cleavage and religation reactions to exchange homologous DNA strands between 34 base-pair (bp) LoxP recognition sequences. In the oligomeric recombination complex, a switch between "cleaving" and "non-cleaving" subunit conformations regulates the number, order, and regio-specificity of the strand exchanges. However, the particular sequence of events has been in question. From analysis of strand composition of the Holliday junction (HJ) intermediate, we determined that Cre initiates recombination of LoxP by cleaving the upper strand on the left arm. Cre preferred to react with the left arm of a LoxP suicide substrate, but at a similar rate to the right arm, indicating that the first strand to be exchanged is selected prior to cleavage. We propose that during complex assembly the cleaving subunit preferentially associates with the LoxP left arm, directing the first strand exchange to that side. In addition, this biased assembly would enforce productive orientation of LoxP sites in the recombination synapses. A novel Cre-HJ complex structure in which LoxP was oriented with the left arm bound by the cleaving Cre subunit suggested a physical basis for the strand exchange order. Lys86 and Lys201 interact with the left arm scissile adenine base differently than in structures that have a scissile guanine. These interactions are associated with positioning the 198-208 loop, a structural component of the conformational switch, in a configuration that is specific to the cleaving conformation. Our results suggest that strand exchange order and site alignment are regulated by an "induced fit" mechanism in which the cleaving conformation is selectively stabilized through protein-DNA interactions with the scissile base on the strand that is cleaved first.

PubMed: 12051940
DOI: 10.1016/S0022-2836(02)00246-2

実験手法

X-RAY DIFFRACTION (2.2 Å)