Reversible protonation of a thiolate ligand in an [Fe]-hydrogenase model complex

Article abstract of DOI:10.1002/anie.201107634

What is the channel? The thiolate ligand in the five-coordinate model complex 1 of [Fe]-hydrogenase is preferentially and reversibly protonated, even in the presence of an acyl ligand. The results suggest that the Cys176 thiolate ligand in [Fe]-hydrogenase can serve as the internal base to accept the proton after heterolytic splitting of H₂.

Full text of DOI:10.1002/anie.201107634

Angewandte
Chemie
DOI: 10.1002/anie.201107634
Biomimetic Chemistry
Reversible Protonation of a Thiolate Ligand in an [Fe]-Hydrogenase
Model Complex**
Dafa Chen, Rosario Scopelliti, and Xile Hu*
[Fe]-hydrogenase is involved in the methanogenic pathway of
the reduction of CO2 to methane.[1–3] In the presence of
methenyltetrahydromethanopterin (methenyl-H4MPT+), this
enzyme catalyzes the heterolytic cleavage of H2 (Scheme 1).[4]
genase. The study indicates that the cysteine thiolate ligand is
a viable proton acceptor for [Fe]-hydrogenase.
According to current spectroscopic[5–12] and crystallo-
graphic data,[13–16] the active site of [Fe]-hydrogenase consists
of a mononuclear FeII complex. The Fe ion is ligated by two
cis-CO, a cysteine sulfur atom (Cys176), and the pyridyl and
acyl donor atoms of a guanylylpyridinol cofactor (Scheme 2).
The coordination site trans to the acyl ligand is proposed as
the H2-binding site. It is unclear whether this site is vacant or
occupied by a H2O molecule in the resting state. Based on this
information, a number of model complexes have been
prepared.[17–31] We recently synthesized a five-coordinate
FeII model complex (1, Scheme 3).[31] This complex provides
a unique platform for reactivity studies, including those of
protonation reactions.
Scheme 1. Enzymatic function of [Fe]-hydrogenase. The reaction has a
DGo’ of À5.5 kJmolÀ1
.
The resulting hydride ion is transferred to methenyl-H4MPT+
to form methylenetetrahydromethanopterin (methylene-
H4MPT). A proton is also formed from this reaction, and it
is shown that this proton exchanges quickly with protons of
the bulk solvent (water).[4] The immediate proton acceptor is
unknown. The cysteine thiolate ligand (Cys176), the pyrido-
nal hydroxy group in the secondary coordination sphere, and
even the acyl ligand of the Fe center (Scheme 2), might
function as the internal base. Herein we describe the
protonation reactions of a model complex of [Fe]-hydro-
Scheme 2. The proposed active site of [Fe]-hydrogenase. The sixth
coordination site might be empty, or occupied by a water molecule.
Cys =cysteine.
Scheme 3. Reactivity of 1 with acids in CH3CN. Py=pyridine.
Reaction of the red compound 1 with HBF4·Et2O in
CH3CN produced a free thiol molecule and a yellow
compound 2 (Scheme 3). The 1H NMR spectrum of 2 exhibits
three signals at 7.98, 7.19 and 6.98 ppm for the pyridyl ring,
two doublets at 4.72 and 3.89 ppm for the diastereotopic
methylene hydrogen atoms, one singlet at 4.01 ppm for the
methoxy group, and one singlet at 1.96 ppm for the coordi-
nated CH3CN molecule.[32] The integration of the signals from
the CH3CN ligands indicates that the Fe center is ligated by
two molecules of CH3CN. In the 13C NMR spectrum of 2, one
acyl carbon (d = 260.6 ppm) and two carbonyl carbon atoms
[*] Dr. D. Chen, Dr. R. Scopelliti, Prof. Dr. X. L. Hu
Laboratory of Inorganic Synthesis and Catalysis
Institute of Chemical Sciences and Engineering
Ecole Polytechnique Fꢀdꢀrale de Lausanne (EPFL)
ISIC-LSCI, BCH 3305, Lausanne 1015 (Switzerland)
E-mail: xile.hu@epfl.ch
Homepage: http://lsci.epfl.ch
[**] This work is supported by the Swiss National Science Foundation
(project no. 200020_134473/1).
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/anie.201107634.
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(d = 208.4 and 207.1 ppm) can be identified. The IR spectrum
of 2 shows two intense absorption bands indicating two cis-
oriented CO ligands (Table 1). Thus, according to the NMR
and IR spectroscopy, complex 2 has the formula of [(2-
acyl ligands. The pyridine ligand sits on the position trans to
one of the CO ligands, which was originally occupied by the
thiolate ligand. The CH3CN ligand occupies the position trans
to the acyl ligand.
The protonation and decoordination of the thiolate ligand
in 1 are reversible. Treatment of 2 and 3 with a mixture of
NEt3 and HS(2,6-Me2C6H3) in CH3CN regenerated complex 1
(Scheme 4). No reaction took place between NEt3 and
Table 1: Selected IR spectroscopy data.
Complex
nCO [cmÀ1
]
1[a]
2
3
2013, 1950
2059, 1997
2042, 1979
2011, 1944
[Fe]-hydrogenase[b]
[a] Data from Ref. [31]. [b] Data from Ref.[6].
CH2CO-6-MeOC5H3N)Fe(CO)2(CH3CN)2]BF4 (Scheme 3).
This formulation is confirmed by elemental analysis.[32]
Unfortunately, we were not able to obtain single crystals of 2.
Reaction of
(PyH·BF4) in CH3CN did not lead to 2, but rather to [(6-
MeOC5H3N-2-CH2CO)Fe(CO)2(CH3CN)(C6H5N)]BF4 (3;
Scheme 3). Compound 3 was also prepared by reaction of 2
with pyridine in CH3CN (Scheme 3). Interestingly, in the
latter reaction, only one CH3CN ligand was replaced by
pyridine, even when an excess amount of pyridine was used.
Single crystals suitable for an X-ray diffraction study were
obtained by diffusion of ether/pentane into a solution of 3 in
CH3CN.
The crystal structure of complex cation in 3 is shown in
Figure 1.[33] The coordination geometry of the Fe center is
octahedral. The acylmethylpyridinyl moiety coordinates by
the pyridyl nitrogen and acyl carbon donors, giving rise to a
five-membered metallacycle. The two CO ligands are cis to
one another; they are both cis to the acyl ligand as well. This
arrangement reflects the strong trans influence of CO and
1
with pyridinium tetrafluoroborate
Scheme 4. Regeneration of 1 from reactions of 2 and 3 with a thiol
ligand under basic conditions.
HS(2,6-Me2C6H3) alone in CH3CN. Thus, HS(2,6-Me2C6H3)
must first coordinate to the Fe ions in 2 and 3 to form Fe-thiol
species as intermediates. Upon binding of sulfur to Fe, the
thiol proton became more acidic and could be deprotonated
by Et3N to give the thiolate complex 1. It is noteworthy that
the thiolate ligand enforces a five-coordinate geometry on the
Fe center, expelling the solvent molecule that originally
occupies the position trans to the acyl ligand. Complex 1 was
also produced by reaction of 2 with 2,6-Me2C6H3SNa.
The reactivity of model compounds 1–3 provides sugges-
tions for the catalytic mechanism of [Fe]-hydrogenase. It
infers that the Cys176 thiolate ligand in the enzyme can be the
immediate proton acceptor after H2 splitting. In the model
system, the resulting thiol molecule is too weak a ligand for
the Fe ion, so it is replaced by a solvent molecule. A similar
ligand substitution reaction may occur in the enzyme after
Cys176 is protonated. It is however possible that the protein
scaffold enforces the sulfur coordination. In any case, the thiol
proton needs to be transferred to a relay base to regenerate
the thiolate ligand, which will restore the active site. This
proton transfer step is fast according to an earlier study.[4] In
the model study, the relay base is Et3N. For the enzyme, the
relay base could be the pyridinol group or the His14 group. It
was reported that a His14 exchange reduced the activity of the
wild enzyme to less than 1%.[16] Speculatively, the important
role of His14 might be to regenerate the thiolate ligand and
thus the active site.
The IR spectra of both 2 and 3 show two intense nCO
absorption bands (2059 and 1997 cmÀ1 for 2; 2042 and
1979 cmÀ1 for 3), consistent with the presence of two cis-CO
groups (Table 1). Compared to complex 1 and [Fe]-hydro-
genase, the averaged nCO stretching frequencies of 2 and 3 are
significantly higher (> 30 cmÀ1). This shift reflects the charge
difference among model compounds 1–3. Cationic Fe com-
plexes in 2 and 3 have significantly higher nCO than the neutral
Figure 1. Solid-state molecular structure of the complex cation in
compound 3. There are two independent molecules in each asymmet-
ric unit; only one of them is shown. Thermal ellipsoids are set at 30%
probability. Selected bond lengths [ꢁ] and angles [o]: Fe1–N1 2.041(6),
Fe1–N2 2.028(6), Fe1–N3 2.028(7), Fe1–C7 1.954(7), Fe1–C14
1.785(8), Fe1–C17 1.770(8), C7–O2 1.200(9), C14–O3 1.150(9), C17–
O4 1.147(9), C14-Fe1-C17 88.6(3).
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Fe complex 1. The IR data in turn suggest that the Fe complex
in the active site of [Fe]-hydrogenase is likely neutral, because
its nCO is very close to that of 1, but not those of 2 and 3
(Table 1).
In summary, the thiolate ligand in complex 1, a unique
small-molecule mimic of the active site of [Fe]-hydrogenase,
can be reversibly protonated. This study suggests that the
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4
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Keywords: enzyme models · hydrogenases · iron · protonation ·
.
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