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Authors
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K.Katayanagi,
M.Miyagawa,
M.Matsushima,
M.Ishikawa,
S.Kanaya,
H.Nakamura,
M.Ikehara,
T.Matsuzaki,
K.Morikawa.
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The crystal structure of RNase H from Escherichia coli has been determined by
the multiple isomorphous replacement method, and refined by the stereochemically
restrained least-squares procedure to a crystallographic R-factor of 0.196 at
1.48 A resolution. In the final structure, the root-mean-square (r.m.s.)
deviation for bond lengths is 0.017 A, and for angle distances 0.036 A. The
structure is composed of a five-stranded beta-sheet and five alpha-helices, and
reveals the details of hydrogen bonding, electrostatic and hydrophobic
interactions between intra- and intermolecular residues. The refined structure
allows an explanation of the particular interactions between the basic
protrusion, consisting of helix alpha III and the following loop, and the
remaining major domain. The beta-sheet, alpha II, alpha III and alpha IV form a
central hydrophobic cleft that contains all six tryptophan residues, and
presumably serves to fix the orientation of the basic protrusion. Two parallel
adjacent helices, alpha I and alpha IV, are associated with a few triads of
hydrophobic interactions, including many leucine residues, that are similar to
the repeated leucine motif. The well-defined electron density map allows
detailed discussion of amino acid residues likely to be involved in binding a
DNA/RNA hybrid, and construction of a putative model of the enzyme complexed
with a DNA/RNA hybrid oligomer. In this model, a protein region, from the
Mg(2+)-binding site to the basic protrusion, covers roughly two turns of a
DNA/RNA hybrid double helix. A segment (11-23) containing six glycine residues
forms a long loop between the beta A and beta B strands. This loop, which
protrudes into the solvent region, lies on the interface between the enzyme and
a DNA/RNA hybrid in the model of the complex. The mean temperature factors of
main-chain atoms show remarkably high values in helix alpha III that constitutes
the basic protrusion, suggesting some correlation between its flexibility and
the nucleic acid binding function. The Mg(2+)-binding site, surrounded by four
invariant acidic residues, can now be described more precisely in conjunction
with the catalytic activity. The arrangement of molecules within the crystal
appears to be dominated by the cancelling out of a remarkably biased charge
distribution on the molecular surface, which is derived in particular from the
separation between the acidic Mg(2+)-binding site and the basic protrusion.
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