Biomimetic intradiol-cleavage of catechols with incorporation of both atoms of O2: The role of the vacant coordination site on the iron center

Article abstract of DOI:10.1246/cl.2001.1062

This is the first example of model system for the active site of protocatechuate 3,4-dioxygenase to display intradiol-cleavage of catechols with incorporation of two oxygen atoms of O₂ promoted by iron complexes.

Full text of DOI:10.1246/cl.2001.1062

1062
Chemistry Letters 2001
Biomimetic Intradiol-Cleavage of Catechols with Incorporation of Both Atoms of O2:
The Role of the Vacant Coordination Site on the Iron Center
Seiji Ogo,*† Ryo Yamahara,†† Takuzo Funabiki,††† Hideki Masuda,†† and Yoshihito Watanabe*†,††††
†Institute for Molecular Science, Myodaiji, Okazaki 444–8585
††Department of Applied Chemistry, Nagoya Institute of Technology, Gokiso-cho, Showa-ku, Nagoya 466-8555
†††Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Sakyo-ku, Kyoto 606-8501
††††Center for Integrative Bioscience, Myodaiji, Okazaki 444-8585
(Received August 9, 2001; CL-010771)
This is the first example of model system for the active site
of protocatechuate 3,4-dioxygenase to display intradiol-cleavage
of catechols with incorporation of two oxygen atoms of O2
promoted by iron complexes.
The metabolic conversion of aromatic compounds to
aliphatic compounds is of fundamental importance in biology.
Catechol dioxygenases are mononuclear non-heme iron
enzymes that catalyze the oxygenation of catechols to aliphatic
acids via cleavage of aromatic rings.1 These enzymes can be
divided into two types: intradiol-cleaving enzymes which break
the catechol C1–C2 bond, and extradiol-cleaving enzymes
which break the C2–C3 or C1–C6 bond. Since Hayaishi et al.
have revealed that an intradiol-cleaving catechol dioxygenase,
pyrocatechase, catalyzes the oxygenation of catechol to muconic
acid with incorporation of two oxygen atoms of O2 (but not of
H2O),2 the oxygenation mechanisms of catechol dioxygenases
have been studied through investigations of model systems3 as
well as the enzymes themselves.4 However, details of the O2
insertion and aromatic ring-cleavage reactions are not yet
understood. Interestingly, recent crystallographic studies of a
protocatechuic acid (PCA)-bound form of an intradiol-cleaving
catechol dioxygenase, protocatechuate 3,4-dioxygenase (3,4-
PCD)5 have revealed that the iron atom in the active site has
octahedral geometry with PCA, His460, His462, Tyr408, and a
vacant coordination site capable of accommodating an exoge-
nous ligand such as an O2 (Figure 1a). Herein, we report the
first example of model system to display intradiol-cleavage of
catechols with incorporation of two oxygen atoms of O2 pro-
moted by iron complexes (Figure 1b): [FeIII(3L)(DBC)Cl]
3
(PPh4) {1, L = N-(2-hydroxyphenyl)-N-(2-pyridylmethyl)ben-
zylamine, DBC = 3,5-di-tert-butylcatecholato} and
[FeIII(3L)(DBC)(DMF)] (2, DMF = N,N-dimethylformamide).
The Cl– and DMF ligands of 1 and 2 are expected to be
exchanged for incoming O2 during the oxygenation.
The new tridentate ligand 3L was designed and synthesized
to mimic specific attributes of the iron coordination site in the
PCA-bound form of 3,4-PCD. Complex 1 was synthesized
tion mode for the three O atoms analogous to the PCA-bound
form of 3,4-PCD. The negative-ion ESI (electrospray ioniza-
tion) mass spectrum of 1 in DMF shows a prominent signal at
m/z 600.2 {relative intensity (I) = 100% in the range of m/z
400–800} which corresponds to [Fe(3L)(DBC)Cl]– ([1]–).
Complex 2 was synthesized from the reaction of 1 with
3
from the reaction of FeCl3 with L, DBC, triethylamine, and
PPh4Cl in DMF. The structure of 1 was unequivocally deter-
mined by X-ray crystallographic analysis.6 An ORTEP draw-
ing of the anion of 1 is shown in Figure 1c. Complex 1 has a
distorted octahedral coordination geometry with bonding
parameters similar to those of the PCA-bound form of 3,4-
PCD.5 Complex 1 has a trans arrangement of O1(phenolato
group) and O2(DBC) atoms and a cis arrangement of O1(phe-
nolato group) and O3(DBC) atoms, i.e., a meridional coordina-
–
AgOTf (OTf = CF3SO3 ) in DMF. The positive-ion ESI mass
spectrum of 2 shows a prominent signal at m/z 639.4 (I = 100%
in the range of m/z 400–800) which corresponds to
[Fe(3L)(DBC)(DMF)+H]+ ([2+H]+). To establish the existence
Copyright © 2001 The Chemical Society of Japan

Chemistry Letters 2001
1063
of the DMF ligand in 2, the same synthesis of 2 has also been
carried out in DMF-d7. ESI-MS results show that the signal at
m/z 639.4 shifts to m/z 646.4 {[Fe(3L)(DBC)(DMF-d7) +H]+},
i.e., the labeled DMF is incorporated into 2.
The oxygenation ability of 1 and 2 in DMF at 25 °C is
shown in Table 1. Complex 1 reacts with O2 to yield intradiol-
products 5 (18% yield based on DBC) and 3,5-di-tert-butyl-5-
(N,N-dimethylamidomethyl)-2-furanone (6, 34% yield) whose
structure was determined by X-ray analysis7 and a non-intradiol-
cleavage product 3,5-di-tert-butyl-1,2-benzoquinone (7, 45%
yield). Complex 2 reacts with O2 to yield 5 (27% yield) and 6
(70% yield). The negative-ion ESI mass spectra of 5 show that
two oxygen atoms of 18O2 are incorporated into 5 upon the
reaction of 1 or 2 with 18O2.
The UV–vis spectra of 1 and 2 in DMF show two features
{1: λmax = 708 nm (ε = 3460 M–1cm–1, M = mol L–1) and 464
(3660) and 2: λmax = 710 (3990) and 466 (4260)} which are
assigned to ligand-to-metal charge transfer (LMCT) transitions
by analogy to those observed in the spectra of other catechol
bound complexes.3 EPR experiments of 1 and 2 observed in
DMF at 77 K indicate that they are high-spin ferric complexes.
We recently reported on the oxygenation ability of a catechol-
bound iron(III) complex with a tetradentate ligand, [FeIII(4L)(DBC)]
{3, 4L = 2-hydroxyphenyl-bis(2-pyridylmethyl)amine}.3a In DMF
at 25 °C, complex 3 reacts with O2 to yield intradiol-cleavage prod-
ucts, 3,5-di-tert-butyl-1-oxacyclohepta-3,5-diene-2,7-dione (4, 73%
yield based on DBC, Table 1) and 3,5-di-tert-butyl-5-(car-
boxymethyl)-2-furanone (5, 26%).3a GC–MS and ESI-MS meas-
urements show that only one oxygen atom of 18O2 is incorporated
into 4 and 5 upon the reaction of 3 with 18O2. The hydrolysis of 4
eventually affords 5 containing one 18O atom.
Furthermore, the kinetic study was followed by monitoring
the disappearance of the lower energy LMCT bands {λmax
=
708 nm (for 1) or 710 (for 2) in DMF}. The reaction rates (kO
2
= kobs/[O2] in DMF at 25 °C)8 of 1 and 2 are 1.80(8) × 10-2 and
2.15(9) × 10–2 M–1s–1, respectively. The oxygenation reactions
(decay of 1 and 2) exhibit pseudo-first-order kinetics.
In summary, what makes 1 and 2 different from 3 is that
the intradiol-cleavage product 5 derived from 1 and 2 is shown
to incorporate both atoms of O2. However, the 18O-labeling
experiments of 3 show that only one label is found in 5. Thus,
depending on the ligands used, either one or two oxygen
atom(s) of O2 are incorporated into the cleavage product. We
attribute these results to the presence and absence of the O2
binding site in these complexes.
Financial support of this research by the Ministry of
Education, Science, Sports, and Culture, Japan Society for the
Promotion of Science, Grant-in-Aid for Scientific Research to
S.O. (13640568) and Y.W. (11490036 and 11228208) is grateful-
ly acknowledged.
References and Notes
1
T. Funabiki, in “Oxygenases and Model Systems, Catalysis by
Metal Complexes” ed. by T. Funabiki, Kluwer, Dordrecht (1997),
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2
3
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4
5
6
A. M. Orville, J. D. Lipscomb, and D. H. Ohlendorf, Biochemistry,
36, 10052 (1997).
Crystallographic data for 1: C57H57ClFeN2O3P, Mr = 940.36, triclin-
–
ic, space group P1 (No. 2), a = 9.7300(9), b = 14.0200(5), c =
18.260(2) Å, α = 78.040(2), β = 78.300(1), γ = 86.950(1)°, V =
2386.1(4) Å3, Z = 2, ρcalcd = 1.309 g cm–3, µ(Mo Kα) = 4.53 cm–1,
R = 0.042 and Rw = 0.107, 10573 reflections used, 814 parameters.
Crystallographic data for 1 have been deposited with the Cambridge
Crystallographic Data Center as supplementary publication no.
CCDC-153553.
7
8
The details of the crystal structure of 6 will be reported elsewhere in
a full paper.
The solubility of O2 (1 atm) in DMF at 25 °C is 4.86 mM. Japan
Chemical Society, Kagaku-Binran Basic Part II, 2nd edition,
Maruzen, Tokyo, 775 (1975).