Modeling of the 5′-deiodination of thyroxine by iodothyronine deiodinase: Chemical corroboration of a selenenyl iodide intermediate

Article abstract of DOI:10.1002/anie.200905796

(Figure Presented) What makes a good cavity? A molecular cavity enabled the stabilization of a selenenyl iodide (RSel) intermediate formed in 5'-deiodination of a thyroxine derivative by an organoselenol (see scheme). The chemical processes proposed for the iodothyronine deiodinase catalytic cycle were experimentally established.

Full text of DOI:10.1002/anie.200905796

Angewandte
Chemie
DOI: 10.1002/anie.200905796
Enzyme Models
Modeling of the 5’-Deiodination of Thyroxine by Iodothyronine
Deiodinase: Chemical Corroboration of a Selenenyl Iodide
Intermediate**
Kei Goto,* Daiju Sonoda, Keiichi Shimada, Shohei Sase, and Takayuki Kawashima*
Type I iodothyronine deiodinase (ID-1) is an enzyme that
catalyzes the conversion of a human thyroid prohormone
(thyroxine; T4) into a biologically active hormone (3,5,3’-
triiodothyronine; T3) through 5’-deiodination (Scheme 1),
and contains a selenocysteine residue at its active site.[1] The
mechanism proposed for the catalytic cycle of ID-1 involves a
ping-pong bisubstrate reaction in which the selenol form of
the enzyme (ESeH) reacts with T4 to form a selenenyl iodide
intermediate (ESeI) with release of the deiodinated com-
pound T3, and a subsequent reaction between the selenenyl
iodide intermediate and possibly a thiol cofactor regenerates
the selenol form.[1,2] The involvement of a selenenyl iodide as
an intermediate in this deiodination process has been widely
accepted, and many model studies on the catalytic cycle of
ID-1 have been performed assuming the formation of such
intermediates.[3] Investigations of antithyroid drugs have been
based on the proposed mechanism depicted in Scheme 1.[4]
However, the chemical evidence for the formation of a
selenenyl iodide in the deiodination reaction of 2,6-diiodo-
phenol derivatives by an organoselenol [Eq. (1)] has been
entirely circumstantial. Although there have been several
reports of the reaction of 2,6-diiodophenol derivatives with
organoselenols,[3a,b] the formation of the corresponding sele-
nenyl iodide intermediates has never been detected. The
verification of the process in Equation (1) has been difficult
not only because of the instability of selenenyl iodides,[5] but
also as a result of the high reactivity of selenenyl iodides
toward selenols.[3c] Herein, we report experimental evidence
for the formation of a selenenyl iodide in 5’-deiodination of a
thyroxine derivative by an organoselenol, through the use of a
nanosized molecular cavity to stabilize the selenenyl iodide
intermediate.
Selenenyl iodides usually undergo facile disproportiona-
tion to the corresponding diselenide and iodine [Eq. (2)].[5] To
demonstrate the process in Equation (1), it is essential that
the selenenyl iodide formed in the reaction not undergo such
disproportionation or react with the parent selenol to produce
the diselenide [Eq. (3)]. There have been several reports of
the isolation of selenenyl iodides stabilized against dispro-
portionation through the introduction of a sterically demand-
ing alkyl substituent (1)[6] or an intramolecular coordinating
group (e.g. 2).[7] However, it has been reported that even 2
reacts with the corresponding selenol to produce the sym-
metrical diselenide within several minutes at room temper-
ature.[3c] In the course of our study on the nanosized molecular
cavities,[8] we previously developed a novel steric protection
group, a Bpq group (cavity-shaped substituent; Scheme 2),[9a]
and showed that it very effectively stabilizes organoselenium
species, such as a Se-nitrososelenol (RSeNO), which are
Scheme 1. Proposed mechanism for the deiodination of thyroxine T4
by ID-1. E=enzyme.
[*] Dr. K. Goto, Dr. S. Sase
Interactive Research Center of Science
Graduate School of Science and Engineering
Tokyo Institute of Technology
2-12-1 Ookayama, Meguro-ku, Tokyo 152-8551 (Japan)
Fax: (+81)3-5734-3543
E-mail: goto@chem.titech.ac.jp
D. Sonoda, Dr. K. Shimada, Prof. Dr. T. Kawashima
Department of Chemistry, Graduate School of Science
The University of Tokyo
7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan)
[**] This work was supported by the Global COE Program (Chemistry)
and Grants-in-Aid for Scientific Research from the Ministry of
Education, Culture, Sports, Science and Technology (Japan) and the
Japan Society for the Promotion of Science. K.G. is grateful to Toray
Science Foundation and The Society of Iodine Science for financial
support. We also thank the Tosoh Finechem Corporation for the
generous gifts of alkyllithium reagents.
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/anie.200905796.
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Scheme 2. Synthesis of a stable selenenyl iodide bearing a cavity-
shaped steric protection group (Bpq).
Scheme 3. Deiodination of the thyroxine derivative 6 by selenol 3. The
yields were estimated by 1H NMR spectroscopy.
otherwise extremely labile.[9b,c] Selenenyl iodide 4 bearing the
Bpq group was synthesized by the reaction of selenol 3[9b] with
N-iodosuccinimide (NIS) and gave a quantitative yield
(Scheme 2). Its structure was established by X-ray crystallog-
raphy (Figure 1).[10] Selenenyl iodide 4 showed remarkable
stability; no decomposition was observed in [D8]toluene after
Similar to the reaction of 6, the reaction of 8 with selenol 3 in
the presence of triethylamine produced monoiodophenol 9
and selenenyl iodide 4 (Scheme 4). However, in the absence
of amine, no reaction took place under similar conditions. In
the active site of ID-1 there are histidine residues near the
selenocysteine catalytic center, and it has been proposed that
the imidazole unit deprotonates the selenol to increase its
reactivity.[11] The present results indicate that the efficiency of
the deiodination process largely depends on the nucleophi-
licity of the selenol functionality.
It has been reported that replacement of the selenocys-
teine residue of ID-1 with cysteine significantly reduces the
catalytic activity of the enzyme.[2a] When thiol 10 bearing a
Bpq group was used instead of selenol 3, no reaction with 8
was observed (Scheme 4). This outcome suggests that the
selenol functionality, with greater nucleophilicity, is essential
for this deiodination reaction.
As a mechanism for the deiodination process shown in
Equation (1), nucleophilic attack of a selenol (or selenolate)
on the iodine center of the keto form of the diiodophenols has
been proposed (Scheme 5).[3b] Consistent with this mecha-
nism, methyl ether 11, which cannot undergo tautomerization
to the keto form, gave no reaction with 3 under these
conditions (Scheme 6). Selective deiodination at the outer
ring of the thyroxine derivative 6 and the inertness of its inner
ring (Scheme 3) are also consistent with the mechanism
involving tautomerization.
1
heating at 1008C in a sealed tube for seven days. In H and
77Se NMR spectra and a UV/Vis spectrum measured at room
temperature, an equilibrium between selenenyl iodide 4 and
the corresponding diselenide was not observed. Treatment of
selenenyl iodide 4 with selenol 3 in CDCl3 at room temper-
ature for seven days resulted in no change. Very slow
formation of the corresponding diselenide, BpqSeSeBpq
(5),[9c] was observed in the presence of triethylamine, but
the yield was only 22% after seven days. These results
indicate that selenenyl iodide 4 has high compatibility with
selenol 3, which is required to demonstrate the process
described by Equation (1). As shown in Figure 1, the SeI
functionality of 4 was incorporated in the cavity, and the
bimolecular processes shown in Equations (2) and (3) are
considered to be effectively prevented by steric repulsion
between the periphery of the molecules.
Deiodination of a thyroxine derivative, N-butyrylthyrox-
ine methyl ester (6),[3b] by selenol 3 was investigated
(Scheme 3). Deiodination of 6 proceeded slowly when it
was treated with an equimolar amount of 3 in the presence of
triethylamine, and after seven days 65% had been converted
into the monodeiodinated product 7 with the concomitant
formation of selenenyl iodide 4 (55% from 3). This is the first
experimental demonstration of the chemical transformation
shown in Equation (1). Notably, only the iodine atom on the
outer phenol ring of 6 was removed; no deiodination at the
inner ring was detected. This result is similar to the conversion
of T4 into T3 in the ID-1 catalytic cycle (Scheme 1).
Control experiments were carried out using 2,6-diiodo-4-
phenoxyphenol (8)[3b] as a model compound of thyroxine.
Scheme 4. Reactions of diiodophenol 8 with selenol 3 or thiol 10. The
yields were estimated by 1H NMR spectroscopy.
Scheme 5. Proposed mechanism for the deiodination process via enol/
keto tautomerization.
Figure 1. The thermal ellipsoids are drawn at the 50% probability level
for 4. Selected bond lengths [ꢀ] and bond angle [8]: I1–Se1 2.5203(11),
Se1–C1 1.946(5); I1-Se1-C1 102.14(15).
Scheme 6. Reaction of methyl ether 11 with selenol 3.
ꢀ 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2010, 49, 545 –547
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c) G. W. Lian, L. Ding, M. Chen, Z. Liu, D. Q. Zhao, J. Z. Ni,
J. Biol. Chem. 2001, 276, 28037 – 28041.
[3] a) C. Beck, S. B. Jensen, J. Reglinski, Bioorg. Med. Chem. Lett.
1994, 4, 1353 – 1356; b) A. A. Vasilꢂev, L. Engman, J. Org. Chem.
1998, 63, 3911 – 3917; c) G. Mugesh, W.-W. du Mont, C. Wism-
ach, P. G. Jones, ChemBioChem 2002, 3, 440 – 447.
Scheme 7. Reduction of selenenyl iodide 4 to selenol 3 by DTT.
[4] a) W.-W. du Mont, G. Mugesh, C. Wismach, P. G. Jones, Angew.
Chem. 2001, 113, 2547 – 2550; Angew. Chem. Int. Ed. 2001, 40,
2486 – 2489; b) G. Roy, M. Nethaji, G. Mugesh, J. Am. Chem.
Soc. 2004, 126, 2712 – 2713; c) G. Roy, G. Mugesh, J. Am. Chem.
Soc. 2005, 127, 15207 – 15217.
[5] a) W.-W. du Mont, A. Martens-von Salzen, F. Ruthe, E. Seppala,
G. Mugesh, F. A. Devillanova, V. Lippolis, N. Kuhn, J. Organo-
met. Chem. 2001, 623, 14 – 28; b) W.-W. du Mont, Main Group
Chem. News 1994, 2, 18 – 26; c) A. Martens-von Salzen, H. U.
Meyer, W.-W. du Mont, Phosphorus Sulfur Silicon Relat. Elem.
1992, 67, 67 – 71; d) W.-W. du Mont, A. Martens, S. Pohl, W.
Saak, Inorg. Chem. 1990, 29, 4847 – 4848; e) W.-W. du Mont, S.
Kubiniok, K. Peters, H. G. Vonschnering, Angew. Chem. 1987,
99, 820 – 821; Angew. Chem. Int. Ed. Engl. 1987, 26, 780 – 781.
[6] a) W.-W. du Mont, I. Wagner, Chem. Ber. 1988, 121, 2109 – 2110;
b) M. Ostrowski, I. Wagner, W.-W. du Mont, P. G. Jones, J. Jeske,
Z. Anorg. Allg. Chem. 1993, 619, 1693 – 1698.
Although the physiological cofactor in the reduction of
the selenenyl iodide intermediate to the selenol in the ID-1
catalytic cycle (Scheme 1) has not been identified, the use of
dithiols, including dithiothreitol (DTT), as the second sub-
strate is common for in vitro experiments with the enzyme.[1b]
Treatment of selenenyl iodide 4 with DTT in the molar ratio
of 1:2.4 in the presence of triethylamine produced selenol 3
quantitatively within 20 minutes (Scheme 7).[12]
In summary, the formation of a selenenyl iodide in the
deiodination of a thyroxine derivative by an organoselenol
was demonstrated for the first time by using cavity-shaped
molecules. In conjunction with reduction of the selenenyl
iodide to the parent selenol by a dithiol, all the chemical
transformations included in the ID-1 catalytic cycle were
experimentally established, thus corroborating the involve-
ment of a selenenyl iodide as an intermediate in the
enzymatic reaction. Further investigations on the mechanistic
details of the processes are currently underway.
[7] a) G. Mugesh, A. Panda, H. B. Singh, R. J. Butcher, Chem. Eur.
J. 1999, 5, 1411 – 1421; b) A. Panda, G. Mugesh, H. B. Singh, R. J.
Butcher, Organometallics 1999, 18, 1986 – 1993; c) G. Mugesh,
H. B. Singh, R. J. Butcher, Tetrahedron: Asymmetry 1999, 10,
237 – 242.
[8] a) K. Goto, N. Tokitoh, R. Okazaki, Angew. Chem. 1995, 107,
1202 – 1203; Angew. Chem. Int. Ed. Engl. 1995, 34, 1124 – 1126;
b) K. Goto, M. Holler, R. Okazaki, J. Am. Chem. Soc. 1997, 119,
1460 – 1461; c) T. Saiki, K. Goto, R. Okazaki, Angew. Chem.
1997, 109, 2320 – 2322; Angew. Chem. Int. Ed. Engl. 1997, 36,
2223 – 2224; d) Y. Ohzu, K. Goto, T. Kawashima, Angew. Chem.
2003, 115, 5892 – 5895; Angew. Chem. Int. Ed. 2003, 42, 5714 –
5717; e) M. Yamashita, K. Goto, T. Kawashima, J. Am. Chem.
Soc. 2005, 127, 7294 – 7295; f) Y.-L. Zhao, M. D. Bartberger, K.
Goto, K. Shimada, T. Kawashima, K. N. Houk, J. Am. Chem.
Soc. 2005, 127, 7964 – 7965.
Experimental Section
Synthesis of selenenyl iodide 4: CCl4 (10 mL) was added to a mixture
of selenol 3 (1.10 g, 1.15 mmol) and N-iodosuccinimide (561 mg,
2.49 mmol) at room temperature. The reaction mixture was stirred for
2 h, filtered through Celite, and the solvent was evaporated.
Recrystallization from n-hexane gave selenenyl iodide 4 (1.23 g,
1.14 mmol, 99%) as purple crystals. 4: m.p. 278.0–281.08C (decomp);
1H NMR (500 MHz, CDCl3): d = 1.07 (d, J = 6.8 Hz, 24H), 1.13 (d,
J = 6.8 Hz, 24H), 2.91 (sept, J = 6.8 Hz, 8H), 7.02 (t, J = 1.5 Hz, 2H),
7.17 (d, J = 7.7 Hz, 8H), 7.24 (d, J = 1.5 Hz, 4H), 7.30 (t, J = 7.7 Hz,
4H), 7.39–7.49 ppm (m, 3H); 13C NMR (126 MHz, [D6]benzene; s, d,
and q are the multiplicities of the signals in the non-decoupled
spectrum): d = 24.1 (q), 24.5 (q), 30.6 (d), 122.7 (d) ꢀ 2, 128.1 (d), 128.3
(d), 129.2 (d), 129.7 (d), 139.2 (s), 140.5 (s), 142.6 (s), 146.8 (s) ꢀ 2,
149.8 ppm (s); 77Se NMR (95 MHz, CDCl3): d = 465 ppm; UV/Vis
(benzene) lmax 553 nm (e 280); elemental analysis calcd (%) for
C66H77ISe: C 73.66, H 7.21; found: C 73.48, H 7.32. For details of the
reaction of 3 with iodophenols, see the Supporting Information.
[9] a) K. Goto, G. Yamamoto, B. Tan, R. Okazaki, Tetrahedron Lett.
2001, 42, 4875 – 4877; b) K. Shimada, K. Goto, T. Kawashima, N.
Takagi, Y.-K. Choe, S. Nagase, J. Am. Chem. Soc. 2004, 126,
13238 – 13239; c) K. Shimada, K. Goto, T. Kawashima, Chem.
Lett. 2005, 34, 654 – 655.
ꢀ
[10] 4·2CHCl3: C68H79Cl6ISe, Mr = 1314.87, triclinic, space group P1,
a = 14.499(7), b = 15.452(7), c = 16.293(6) ꢃ, a = 69.028(12), b =
73.775(11), g = 88.047(15)8, V= 3264(2) ꢃ3, Z = 2, 1calcd
=
1.338 gcmÀ3, T= 120 K, m(MoKa) = 1.332 mmÀ1, 21498 measured
reflections, 11269 independent, 729 parameters, R1 = 0.0635 (I >
2s(I)), wR2 = 0.1781 (all data). The intensity data were collected
on a Rigaku/MSC Mercury CCD diffractometer with graphite-
monochromated MoKa radiation (l = 0.71070 ꢃ). The structures
were solved by the direct method and refined by full-matrix least
squares on F2 using SHELXL 97 (G. M. Sheldrick, Program for
Crystal Structure Refinement, University of Gꢁttingen, 1997).
The non-hydrogen atoms were refined anisotropically. The
hydrogen atoms were idealized by using the riding models.
CCDC 750141 contains the supplementary crystallographic data
for this paper. These data can be obtained free of charge from
The Cambridge Crystallographic Data Centre via www.ccdc.
cam.ac.uk/data_request/cif.
Received: October 15, 2009
Published online: December 8, 2009
Keywords: enzyme models · iodine · reactive intermediates ·
.
selenium · steric hindrance
[1] a) M. J. Berry, L. Banu, P. R. Larsen, Nature 1991, 349, 438 – 440;
b) P. R. Larsen, M. J. Berry, Annu. Rev. Nutr. 1995, 15, 323 – 352;
c) A. C. Bianco, D. Salvatore, B. Gereben, M. J. Berry, P. R.
Larsen, Endocr. Rev. 2002, 23, 38 – 89; d) J. Kꢁhrle, Methods
Enzymol. 2002, 347, 125 – 167; e) G. Kuiper, M. H. A. Kester,
R. P. Peeters, T. J. Visser, Thyroid 2005, 15, 787 – 798; f) D. L. St.
Germain, V. A. Galton, A. Hernandez, Endocrinology 2009, 150,
1097 – 1107.
[11] M. J. Berry, J. Biol. Chem. 1992, 267, 18055 – 18059.
[12] The reduction of selenenyl iodide 2 has been investigated by
du Mont and co-workers.[3c] It was found that the reaction of 2
with monothiols proceeds readily to afford the corresponding
selenenyl sulfides (ArSeSR) although further reduction to the
selenol is very slow.
[2] a) M. J. Berry, J. D. Kieffer, J. W. Harney, P. R. Larsen, J. Biol.
Chem. 1991, 266, 14155 – 14158; b) B. C. Sun, J. W. Harney, M. J.
Berry, P. R. Larsen, Endocrinology 1997, 138, 5452 – 5458;
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