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The structures of the cyanide and triiodide complexes of Arthromyces ramosus
peroxidase (ARP) at different pH values were investigated by x-ray
crystallography in order to examine the behavior of the invariant residues of
arginine (Arg-52) and distal histidine (His-56) during the enzyme reaction as
well as to provide the structural basis of the active site of peroxidase. The
models of the cyanide complexes at pH 7.5, 5.0, and 4.0, respectively, were
refined to the R-factors of 17.8, 17.8, and 18.5% using 7.0-1.6-A resolution
data, and those of the triiodide complexes at pH 6.5 and 5.0 refined to 16.9 and
16.8% using 7.0-1.9-A resolution data. The structures of the cyanide complexes
at pH 7.5, 5.0, and 4.0 are identical within experimental error. Cyanide ion
bound to the heme in the bent conformation rather than in the tilt conformation.
Upon cyanide ion binding, the N epsilon atom of His-56 moved toward the ion by
rotation of the imidazole ring around the C beta-C gamma bond, but there was
little conformational change in the remaining residues. The distance between the
N epsilon atom of His-56 and the nitrogen atom of the cyanide suggests the
presence of a hydrogen bond between them in the pH range investigated. In the
triiodide complexes, one of the two triiodides bound to ARP was located at the
distal side of the heme. When triiodide bound to ARP, unlike the rearrangement
of the distal arginine of cytochrome c peroxidase that occurs on formation of
the fluoride complex or compound I, the side chain of Arg-52 moved little. The
conformation of the side chain of His-56, however, changed markedly.
Conformational flexibility of His-56 appears to be a requisite for proton
translocation from one oxygen atom to the other of HOO- by acid-base catalysis
to produce compound I. The iron atom in each cyanide complex (low-spin ferric)
is located in the heme plane, whereas in each triiodide complex (high-spin
ferric) the iron atom is displaced from the plane about 0.2 A toward the
proximal side.
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