5E63

K262A mutant of I-SmaMI

5E63 の概要

• エントリーDOI: 10.2210/pdb5e63/pdb
• 分子名称: I-SmaMI LAGLIDADG meganuclease, DNA right half site Bottom strand, DNA left half site bottom strand, ... (9 entities in total)
• 機能のキーワード: laglidadg, i-smami, k262a, mutant, hydrolase-dna complex, hydrolase/dna
• 由来する生物種: Sordaria macrospora (strain ATCC MYA-333 / DSM 997 / K(L3346) / K-hell); 詳細
• タンパク質・核酸の鎖数: 5
• 化学式量合計: 49630.71

構造登録者

Shen, B.,Stoddard, B. (登録日: 2015-10-09, 公開日: 2016-01-13, 最終更新日: 2023-09-27)

主引用文献

Shen, B.W.,Lambert, A.,Walker, B.C.,Stoddard, B.L.,Kaiser, B.K.
The Structural Basis of Asymmetry in DNA Binding and Cleavage as Exhibited by the I-SmaMI LAGLIDADG Meganuclease.
J.Mol.Biol., 428:206-220, 2016

PubMed Abstract: LAGLIDADG homing endonucleases ("meganucleases") are highly specific DNA cleaving enzymes that are used for genome engineering. Like other enzymes that act on DNA targets, meganucleases often display binding affinities and cleavage activities that are dominated by one protein domain. To decipher the underlying mechanism of asymmetric DNA recognition and catalysis, we identified and characterized a new monomeric meganuclease (I-SmaMI), which belongs to a superfamily of homologous enzymes that recognize divergent DNA sequences. We solved a series of crystal structures of the enzyme-DNA complex representing a progression of sequential reaction states, and we compared the structural rearrangements and surface potential distributions within each protein domain against their relative contribution to binding affinity. We then determined the effects of equivalent point mutations in each of the two enzyme active sites to determine whether asymmetry in DNA recognition is translated into corresponding asymmetry in DNA cleavage activity. These experiments demonstrate the structural basis for "dominance" by one protein domain over the other and provide insights into this enzyme's conformational switch from a nonspecific search mode to a more specific recognition mode.

PubMed: 26705195
DOI: 10.1016/j.jmb.2015.12.005

実験手法

X-RAY DIFFRACTION (2.6 Å)