1EVX: APO CRYSTAL STRUCTURE OF THE HOMING ENDONUCLEASE, I-PPOI

Citation:
Abstract
The homing endonuclease I-PpoI severely bends its DNA target, resulting in significant deformations of the minor and major groove near the scissile phosphate groups. To study the role of conformational changes within the protein catalyst and the DNA substrate, we have determined the structure of the enzyme in the absence of bound DNA, performed gel retardation analyses of DNA binding and bending, and have mutagenized a leucine residue that contacts an adenine nucleotide at the site of cleavage. The structure of the L116A/DNA complex has been determined and the effects of the mutation on affinity and catalysis have been measured. The wild-type protein displays a rigid-body rotation of its individual subunits upon DNA binding. Homing site DNA is not detectably bent in the absence of protein, but is sharply bent in both the wild-type and L116A complexes. These results indicate that binding involves a large distortion of the DNA and a smaller change in protein conformation. Leucine 116 is critical for binding and catalysis: it appears to be important for forming a well-ordered protein-DNA complex at the cleavage site, for maximal deformation of the DNA, and for desolvation of the nucleotide bases that are partially unstacked in the enzyme complex.
PDB ID: 1EVXDownload
MMDB ID: 13739
PDB Deposition Date: 2000/4/20
Updated in MMDB: 2011/11
Experimental Method:
x-ray diffraction
Resolution: 2  Å
Source Organism:
Similar Structures:
Biological Unit for 1EVX: dimeric; determined by author and by software (PISA)
Molecular Components in 1EVX
Label Count Molecule
Proteins (2 molecules)
2
Intron-encoded Homing Endonuclease I-ppoi
Molecule annotation
Chemicals (8 molecules)
1
4
2
4
* Click molecule labels to explore molecular sequence information.

Citing MMDB
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