Nucleic Acids Research, Vol 24, Issue 20 3982-3989, Copyright © 1996 by Oxford University Press
J Lykke-Andersen, RA Garrett and J Kjems
The archaeal intron-encoded homing enzymes I-PorI and I-DmoI belong to a
family of endonucleases that contain two copies of a characteristic
LAGLIDADG motif. These endonucleases cleave their intron- or intein-
alleles site-specifically, and thereby facilitate homing of the introns or
inteins which encode them. The protein structure and the mechanism of DNA
recognition of these homing enzymes is largely unknown. Therefore, we
examined these properties of I-PorI and I-DmoI by protein footprinting.
Both proteins were susceptible to proteolytic cleavage within regions that
are equidistant from each of the two LAGLIDADG motifs. When complexed with
their DNA substrates, a characteristic subset of the exposed sites, located
in regions immediately after and 40-60 amino acids after each of the
LAGLIDADG motifs, were protected. Our data suggest that the enzymes are
structured into two, tandemly repeated, domains, each containing both the
LAGLIDADG motif and two putative DNA binding regions. The latter contains a
potentially novel DNA binding motif conserved in archaeal homing enzymes.
The results are consistent with a model where the LAGLIDADG endonucleases
bind to their non-palindromic substrates as monomeric enzymes, with each of
the two domains recognizing one half of the DNA substrate.
ARTICLES
Protein footprinting approach to mapping DNA binding sites of two archaeal homing enzymes: evidence for a two-domain protein structure
Institute of Molecular Biology, Copenhagen University, Denmark.
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