Resonance Raman study of a high-valent Fe=O porphyrin complex as a model for peroxidase compound II

Article abstract of DOI:10.1246/cl.2010.332

Resonance Raman spectroscopy is applied to a Fe^IV oxo porphyrin with imidazolate as the axial ligand. The V_Fe=O mode is observed at 792 cm^-1, which is 23 cm^-1 lower than that of the analogous 1-methylimidazole complex and similar to that of horseradish peroxidase compound II (787 cm^-1) at alkaline pH, for which presence of an anionic histidine was previously postulated. This study thus provides a useful model compound of horseradish peroxidase compound II.

Full text of DOI:10.1246/cl.2010.332

doi:10.1246/cl.2010.332
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Published on the web February 27, 2010
Resonance Raman Study of a High-valent Fe=O Porphyrin Complex
as a Model for Peroxidase Compound II
Hirohito Ishimaru, Hiroshi Fujii, and Takashi Ogura*1
1
2,3
1
Department of Life Science and Picobiology Institute, Graduate School of Life Science, University of Hyogo,
3
-2-1 Koto, Kamigori-cho, Ako-gun, Hyogo 678-1297
2
Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Aza-higashiyama, Myodaiji, Okazaki 444-8787
3
Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences,
5-1 Aza-higashiyama, Myodaiji, Okazaki 444-8787
(
Received January 21, 2010; CL-100067; E-mail: ogura@sci.u-hyogo.ac.jp)
Resonance Raman spectroscopy is applied to a FeIV oxo
porphyrin with imidazolate as the axial ligand. The ¯Fe=O mode
¹
1
¹1
is observed at 792 cm , which is 23 cm lower than that of the
analogous 1­methylimidazole complex and similar to that of
¹
1
horseradish peroxidase compound II (787 cm ) at alkaline pH,
for which presence of an anionic histidine was previously
postulated. This study thus provides a useful model compound
of horseradish peroxidase compound II.
Horseradish peroxidase (HRP) catalyzes the oxidation of
organic substrates using H2O2 as a specific oxidant. Upon
reaction with H2O2, HRP sequentially forms two reaction
intermediates known as compound I and compound II, before
returning to the original ferric state. Compound I and compound
II are respectively 2 and 1 oxidative equivalents higher than the
III
1
V
IV
Fe state, and respectively correspond to the Fe and Fe
formal oxidation states. Compound II of HRP exhibits a ¯Fe=O
¹
1
resonance Raman (RR) band at 775 and 787 cm at pH 7 and
1, respectively. The 12 cm upshift of the ¯Fe=O mode of
2
¹1
1
IV
Figure 1. Absorption spectra of (TMP)(Fe =O) in the absence
HRP compound II upon alkalization is caused by elimination of
and presence of trans ligand at ¹40 °C in chlorobenzene.
the hydrogen bond between the oxygen atom and the distal His
IV
IV
(
A) (TMP)(Fe =O), (B) (TMP)(Fe =O)(1MeIm), and (C)
IV
IV
¹
and is regarded as a “distal effect.” The ¯
mode of the Fe
(TMP)(Fe =O)(Im ). The concentration of TMP was 0.2 mM in
the presence of 0, 0.2, and 0.4 mM ligand for (A), (B), and (C).
Expanded spectra in the visible region are for 1 mM TMP in the
presence of 0, 1, and 2 mM ligand for (A), (B), and (C).
Fe=O
3
¹1
state of myoglobin (Mb) at pH 8.5 is located at ca. 800 cm
(
sperm whale Mb at 797 cm and horse heart Mb at 804 cm¹1),
¹1
where no hydrogen bond exists between the oxygen atom and
¹
1
the distal His. There is a 13 cm frequency difference in the
¹
1
¹1
IV
¯
Fe=O mode between HRP at 787 cm and Mb at 800 cm
.
tion maxima at 416 and 543 nm. (TMP)(Fe =O)(1MeIm)
This has been interpreted as a result of a “proximal effect.” Both
has absorption maxima at 425, 556, 569, and 604 nm.
HRP and Mb have a proximal His residue.4 However, the
,5
(TMP)(Fe =O)(Im ) has absorption maxima at 423, 558,
566, and 604 nm. The absorption spectra of the 6-coordinate
complexes (Figures 1B and 1C) are similar to each other and
distinct from that of the 5-coordinate complex (Figure 1A).
IV
¹
proximal His of HRP has the anionic character of imidazolate
¹
(
Im ), while the proximal His of Mb is considered to be neutral.
Many Fe=O porphyrin model complexes with 1-methyl-
IV 16
imidazole (1MeIm) or a solvent molecule acting as the axial
ligand have been prepared and characterized by RR spectrosco-
Figure 2 depicts RR spectra of (TMP)(Fe = O) (A),
IV 16
IV 16
¹
(TMP)(Fe = O)(1MeIm) (B), and (TMP)(Fe = O)(Im )
(C). Figures 2A¤, 2B¤, and 2C¤ are obtained for the correspond-
6
py, in order to obtain insights into the electronic structures
1
8
and reactivities of hemoproteins. However, there have been no
ing O-complexes and Figures 2A¤¤, 2B¤¤, and 2C¤¤ are the
isotopic difference spectra (see the legend). RR difference
spectra designated by A¤¤, B¤¤, and C¤¤ exhibit the ¯Fe=O mode
at 841/809, 815/778, and 792/760 cm , respectively, for the
O/ O complex. Upon addition of 1MeIm, the ¯Fe=O mode at
841 cm for the 5-coordinate species (Figure 2A) becomes
downshifted to 815 cm (Figure 2B). This downshift is 5 cm
lower than the downshift observed for the same complex in
toluene. Upon addition of Im , instead of 1MeIm, the ¯Fe=O
mode downshifts further to 792 cm by 23 cm (Figure 2C).
Notably the pattern and the frequency of the porphyrin vibra-
¹
reports of model complexes with Im as a trans axial ligand. In
the present study, we have prepared an Fe=O porphyrin model
¹
¹1
complex with Im as the axial ligand and identified the ¯Fe=O
¹
1
16
18
mode at 792 cm , which is significantly lower than that of an
¹
1
¹1
analogous complex with 1MeIm as the axial ligand (815 cm ).
Thus, the imidazolate complex could be regarded as a model for
compound II of HRP. The experimental details are described in
Supporting Information.7
¹
1
¹1
6
¹
8
IV
¹1
¹1
Figure 1 depicts absorption spectra of (TMP) (Fe =O) (A),
¹
IV
+
1MeIm (B), and +Im (C). (TMP)(Fe =O) gives absorp-
Chem. Lett. 2010, 39, 332­333
© 2010 The Chemical Society of Japan
www.csj.jp/journals/chem-lett/

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33
¹
frequency for HRP is also due to the Im character of its
9
4
proximal His. The X-ray structure of HRP revealed the
presence of a hydrogen bond between the NH of the distal
His170 and the main chain C=O of Asp247, which
gives anionic character to His170. For the model com-
9
,11
II
¹ 8
pounds, the ¯Fe­His modes
of Fe (OEP)(2MeIm ) and
Fe (OEP)(2MeIm) are located at 233 and 208 cm¹1, consistent
with the trend identified in the proteins. An inverse relationship
between the ¯Fe=O and the ¯Fe­His frequencies, has been pointed
out. This indicates that the anionic character of the proximal
His is common for the Fe and Fe states.
II
8
6
a
II
IV
¹
In conclusion, the present results demonstrate that the Im
ligand actually lowers the ¯Fe=O frequency relative to that of
IV
¹
neutral 1MeIm ligand and that the (TMP)(Fe =O)(Im ) could
serve as a model complex for compound II of HRP.
Since the ¯Fe=O frequency is identical for complexes of
IV
¹
IV
(
TMP)(Fe =O) + Im and (TMP)(Fe =O) + Im systems, it
is deduced that the NH proton of Im is removed upon ligation to
IV
Fe . The high basicity of the oxygen atom may contribute to the
deprotonation. It is also likely that the high valent state of Fe
significantly lowers the pKa of the NH proton of Im. These
characteristics are expected to be important for proper function-
ing of certain biological systems, since formation of high-valent
iron complexes such as compound II in the protein during
enzyme catalysis reaction could be communicated to the
proximal side via this mechanism.
T. O. acknowledges support from a Grant-in-Aid for
Scientific Research (C) (No. 21570171), Grant-in-Aid for
Scientific Research on Priority Areas (No. 477) and the Global
Center of Excellence Program, “Picobiology” at the University
of Hyogo with support from MEXT, Japan.
IV
Figure 2. RR spectra of (TMP)(Fe =O) in the absence and
presence of trans ligand at ¹40 °C in chlorobenzene. (A)
IV
IV
(
(
TMP)(Fe =O),
(B)
(TMP)(Fe =O)(1MeIm),
and
(C)
IV
¹
TMP)(Fe =O)(Im ). The concentration of TMP was 1 mM. The
concentration of ligand was 0, 1, and 2 mM for (A), (B), and (C).
16
Spectra (A), (B), and (C) were obtained for O-complexes. Spectra
(
A¤), (B¤), and (C¤) for 18O-complexes. Spectra (A¤¤), (B¤¤), and (C¤¤)
References and Notes
16
18
are isotopic difference spectra ( O­ O: A¤¤ = A­A¤, B¤¤ = B­B¤,
and C¤¤ = C­C¤).
1
a) H. B. Dunford, J. S. Stillman, Coord. Chem. Rev. 1976, 19, 187. b)
I. Yamazaki, M. Tamura, R. Nakajima, Mol. Cell. Biochem. 1981, 40,
1
43.
IV
2
a) S. Hashimoto, Y. Tatsuno, T. Kitagawa, Proc. Jpn. Acad., Ser B
tional modes obtained for (TMP)(Fe =O)(1MeIm) and
(
2
1
1
984, 60, 345. b) A. J. Sitter, C. M. Reczek, J. Terner, J. Biol. Chem.
985, 260, 7515. c) S. Hashimoto, Y. Tatsuno, T. Kitagawa, Proc.
IV
¹
TMP)(Fe =O)(Im ) are virtually identical (Figures 2B and
C), indicating that structural differences in the porphyrin core
Natl. Acad. Sci. U.S.A. 1986, 83, 2417. d) W. A. Oertling, G. T.
Babcock, Biochemistry 1988, 27, 3331.
a) J. Terner, A. J. Sitter, C. M. Reczek, Biochim. Biophys. Acta 1985,
are essentially negligible. On the other hand, the difference of
these two complexes is prominent in the ¯Fe=O mode. Since each
of the ¯Fe=O frequencies in Figures 2B and 2C were unchanged
for the ligand concentration between 0.25 and 10 mM (data not
shown), it is not likely that a direct interaction of the added
3
8
28, 73. b) R. K. Behan, M. T. Green, J. Inorg. Biochem. 2006, 100,
4
48. c) K. Ikemura, M. Mukai, H. Shimada, T. Tsukihara, S.
Yamaguchi, K. Shinzawa-Itoh, S. Yoshikawa, T. Ogura, J. Am. Chem.
Soc. 2008, 130, 14384.
ligand with the oxygen atom causes the downshift in the ¯
4
5
6
G. I. Berglund, G. H. Carlsson, A. T. Smith, H. Szoke, A. Henriksen,
J. Hajdu, Nature 2002, 417, 463.
H.-P. Hersleth, B. Dalhus, C. H. Gorbitz, K. K. Andersson, J. Biol.
Inorg. Chem. 2002, 7, 299.
Fe=O
mode. The ¯Fe=O mode of (TMP)(Fe =O)(Im) is at 792 cm¹1,
IV
8
and addition of tetrabutylammonium hydroxide, which is
¹
expected to remove the NH proton to generate Im , does not
a) W. A. Oertling, R. T. Kean, R. Wever, G. T. Babcock, Inorg. Chem.
1990, 29, 2633. b) T. Kitagawa, Y. Mizutani, Coord. Chem. Rev.
1994, 135­136, 685. c) K. Nakamoto, Coord. Chem. Rev. 2002, 226,
change the frequency (data not shown).
Downshift of the ¯Fe=O mode upon transformation from the
1
53. d) J. Terner, V. Palaniappan, A. Gold, R. Weiss, M. M.
5- to 6-coordinate complex by ligation of 1MeIm is caused by
Fitzgerald, A. M. Sullivan, C. M. Hosten, J. Inorg. Biochem. 2006,
00, 480.
electron donation from the nitrogen atom of 1MeIm to Fe, which
reduces donation from the oxo ligand, resulting in the weaken-
1
7
8
Supporting Information is available electronically on the CSJ-Journal
Web site, http://www.csj.jp/journals/chem-lett/index.html.
Abbreviations: Im; imidazole, OEP; octaethylporphyrin dianion,
6
ing of the Fe=O bond. The lower ¯Fe=O frequency of HRP
¹
1
compound II (787 cm ) relative to the Mb ferryl state
¹1
TMP; tetramesitylporphyrin dianion, 2MeIm; 2-methylimidazole,
(
800 cm ) has been interpreted as a result of larger electron
¹
2
MeIm ; 2-methylimidazolate.
¹
donation from His, whereas the proximal His of HRP has Im
9
J. Teraoka, T. Kitagawa, J. Biol. Chem. 1981, 256, 3969.
9
10
II
character. The ¯Fe­His modes of HRP and Mb in the Fe state
10 T. Kitagawa, K. Nagai, M. Tsubaki, FEBS Lett. 1979, 104, 376.
11 P. Stein, M. Mitchell, T. G. Spiro, J. Am. Chem. Soc. 1980, 102, 7795.
¹
1
are located at 244 and 220 cm , respectively, and the higher
Chem. Lett. 2010, 39, 332­333
© 2010 The Chemical Society of Japan
www.csj.jp/journals/chem-lett/