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PDBsum entry 1ccp
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Oxidoreductase
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PDB id
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1ccp
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References listed in PDB file
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Key reference
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Title
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X-Ray structures of recombinant yeast cytochrome c peroxidase and three heme-Cleft mutants prepared by site-Directed mutagenesis.
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Authors
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J.M.Wang,
M.Mauro,
S.L.Edwards,
S.J.Oatley,
L.A.Fishel,
V.A.Ashford,
N.H.Xuong,
J.Kraut.
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Ref.
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Biochemistry, 1990,
29,
7160-7173.
[DOI no: ]
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PubMed id
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Abstract
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The 2.2-A X-ray structure for CCP(MI), a plasmid-encoded form of Saccharomyces
cerevisiae cytochrome c peroxidase (CCP) expressed in Escherichia coli [Fishel,
L.A., Villafranca, J. E., Mauro, J. M., & Kraut, J. (1987) Biochemistry 26,
351-360], has been solved, together with the structures of three specifically
designed single-site heme-cleft mutants. The structure of CCP(MI) was solved by
using molecular replacement methods, since its crystals grow differently from
the crystals of CCP isolated from bakers' yeast used previously for structural
solution. Small distal-side differences between CCP(MI) and bakers' yeast CCP
are observed, presumably due to a strain-specific Thr-53----Ile substitution in
CCP(MI). A Trp-51----Phe mutant remains pentacoordinated and exhibits only minor
distal structural adjustments. The observation of a vacant sixth coordination
site in this structure differs from the results of solution resonance Raman
studies, which predict hexacoordinated high-spin iron [Smulevich, G., Mauro,
J.M., Fishel, L. A., English, A. M., Kraut, J., & Spiro, T. G. (1988)
Biochemistry 27, 5477-5485]. The coordination behavior of this W51F mutant is
apparently altered in the presence of a precipitating agent, 30%
2-methyl-2,4-pentanediol. A proximal Trp-191----Phe mutant that has
substantially diminished enzyme activity and altered magnetic properties [Mauro,
J. M., Fishel, L. F., Hazzard, J. T., Meyer, T. E., Tollin, G., Cusanovich, M.
A., & Kraut, J. (1988) Biochemistry 27, 6243-6256] accommodates the
substitution by allowing the side chain of Phe-191, together with the segment of
backbone to which it is attached, to move toward the heme. This relatively large
(ca. 1 A) local perturbation is accompanied by numerous small adjustments
resulting in a slight overall compression of the enzyme's proximal domain;
however, the iron coordination sphere is essentially unchanged. This structure
rules out a major alteration in protein conformation as a reason for the
dramatically decreased activity of the W191F mutant. Changing proximal Asp-235
to Asn results in two significant localized structural changes. First, the heme
iron moves toward the porphyrin plane, and distal water 595 now clearly resides
in the iron coordination sphere at a distance of 2.0 A. The observation of
hexacoordinated iron for the D235N mutant is in accord with previous resonance
Raman results. Second, the indole side chain of Trp-191 has flipped over as a
result of the mutation; the tryptophan N epsilon takes part in a new hydrogen
bond with the backbone carbonyl oxygen of Leu-177.(ABSTRACT TRUNCATED AT 400
WORDS)
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Secondary reference #1
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Title
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Crystal structure of yeast cytochrome c peroxidase refined at 1.7-A resolution.
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Authors
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B.C.Finzel,
T.L.Poulos,
J.Kraut.
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Ref.
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J Biol Chem, 1984,
259,
13027-13036.
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PubMed id
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