The catalytic domain of lambda site-specific recombinase.
The catalytic domain of lambda site-specific recombinase.
复制标题
lambda 位点特异性重组酶的催化结构域。
DOI：
10.1073/pnas.94.12.6104
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发表时间：
1997
影响因子：
11.1
通讯作者：
Landy,A
中科院分区：
文献类型：
--
作者：
Tirumalai,RS;Healey,E;Landy,A
TheEscherichia coliphage λ integrase protein (Int) belongs to the large Int family of site-specific recombinases. It is a heterobivalent DNA binding protein that makes use of a high energy covalent phosphotyrosine intermediate to catalyze integrative and excisive recombination at specific chromosomal sites (attsites). A 293-amino acid carboxy-terminal fragment of Int (C65) has been cloned, characterized, and used to further dissect the protein. From this we have cloned and characterized a 188-amino acid, protease-resistant, carboxy-terminal fragment (C170) that we believe is the minimal catalytically competent domain of Int. C170 has topoisomerase activity and convertsattsuicide substrates to the covalent phosphotyrosine complexes characteristic of recombination intermediates. However, it does not show efficient binding toattsite DNA in a native gel shift assay. We propose that λ Int consists of three functional and structural domains: residues 1–64 specify recognition of “arm-type” DNA sequences distant from the region of strand exchange; residues 65–169 contribute to specific recognition of “core-type” sequences at the sites of strand exchange and possibly to protein–protein interactions; and residues 170–356 carry out the chemistry of DNA cleavage and ligation. The finding that the active site nucleophile Tyr-342 is in a uniquely protease-sensitive region complements and reinforces the recently solved C170 crystal structure, which places Tyr-342 at the center of a 17-amino acid flexible loop. It is proposed that C170 is likely to represent a generic Int family domain that thus affords a specific route to studying the chemistry of DNA cleavage and ligation in these recombinases.
