Synthesis of 2-amino-3-fluoroacrylic acid containing peptides

Article abstract of DOI:10.1021/ol006997s

(Matrix presented) Peptides containing (E)- and (Z)-3-fluorodehydroalanine have been prepared from serine via a fluoro-Pummerer rearrangement. The resulting electrophilic moieties may be useful affinity labels for the identification of the targets of dehydroamino acid containing natural products that act by covalent mechanisms.

Full text of DOI:10.1021/ol006997s

ORGANIC
LETTERS
2001
Vol. 3, No. 4
593-596
Synthesis of 2-Amino-3-fluoroacrylic
Acid Containing Peptides
Hao Zhou and Wilfred A. van der Donk*
Department of Chemistry, UniVersity of Illinois at UrbanasChampaign,
600 South Mathews AVenue, Urbana, Illinois 61801
Vddonk@uiuc.edu
Received December 13, 2000
ABSTRACT
Peptides containing (E)- and (Z)-3-fluorodehydroalanine have been prepared from serine via a fluoro-Pummerer rearrangement. The resulting
electrophilic moieties may be useful affinity labels for the identification of the targets of dehydroamino acid containing natural products that
act by covalent mechanisms.
Dehydroalanines are present in a large number of natural
products, including the microcystins,1 nodularin,2 thiostrepton
and other thiopeptides,3 and the lantibiotics.4 The precise
role of these R,â-unsaturated amides in the biological
activities of these compounds is not always well understood.
Their function might be strictly structural or they may act
as electrophilic entities forming covalent linkages with
nucleophilic groups on their targets. Michael acceptors such
as the dehydroalanines have been popular functionalities for
the design of enzyme inhibitors and active site affinity
labels.5 The reversibility of conjugate additions, however,
poses a potential drawback for the permanent attachment of
a target to a Michael acceptor, thereby in certain cases
precluding identification of the nucleophile. Introduction of
a halogen on the terminal vinyl carbon can alleviate this
shortcoming since it will lead to an irreversible linkage of
the nucleophile to the dehydroalanine (Scheme 1). This
Scheme 1
(1) (a) Botes, D.; Tuinman, A.; Wessels, P.; Viljoen, C.; Kruger, H.;
Williams, D. H.; Santikarn, S.; Smith, R.; Hammond, S. J. Chem. Soc.,
Perkin Trans. 1 1984, 2311-2318. (b) Painuly, P.; Perez, R.; Fukai, T.;
Shimizu, Y. Tetrahedron Lett. 1988, 29, 11-14. (c) Dawson, R. M. Toxicon
1998, 36, 953-962.
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373-384. (b) Botes, D.; Wessels, P.; Kruger, H.; Runnegar, M. T. C.;
Santikarn, S.; Smith, R.; Barna, J. C. J.; Williams, D. H. J. Chem. Soc.,
Perkin Trans. 1 1985, 2747-2748. (c) Rinehart, K. L.; Harada, K.;
Namikoshi, M.; Chen, C.; Harvis, C.; Munro, M. H. G.; Blunt, J.; Mulligan,
P.; Beasley, V.; Dahlem, A.; Carmicheal, W. J. Am. Chem. Soc. 1988, 110,
8557-8558.
(3) (a) Anderson, B.; Hodgkin, D. C.; Viswamitra, M. A. Nature 1970,
225, 233-5. (b) Walker, J.; Olesker, A.; Valente, L.; Rabanal, R.; Lukacs,
G. J. Chem. Soc. Chem. Commun. 1977, 706-708. (c) Bycroft, B. W.;
Gowland, M. S. J. Chem. Soc. Chem. Commun. 1978, 256-258. (d) Lau,
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Prod. Rep. 1999, 16, 575-87. (c) Jack, R. W.; Jung, G. Curr. Opin. Chem.
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strategy has been very successful for instance in the study
of pyridoxal phosphate dependent enzymes5,6 as well as
(5) (a) Walsh, C. T. Tetrahedron 1982, 38, 871-909. (b) Walsh, C. T.
Annu. ReV. Biochem. 1984, 53, 493-535. (c) Silverman, R. B. Mechanism-
based enzyme inactiVation: chemistry and enzymology; CRC Press: Boca
Raton, FL, 1988.
10.1021/ol006997s CCC: $20.00 © 2001 American Chemical Society
Published on Web 01/31/2001

D-xylose isomerase,7 and monoamine oxidase.8 Fluorine-
substituted Michael acceptors have been particularly attrac-
tive because of the small steric requirements of the fluorine
substituent. To date, no reports have detailed the preparation
of fluorine-substituted dehydroalanines in peptides. We report
here the synthesis of 3-fluorodehydroalanine-containing
peptides 2.
A number of methods have been reported for the prepara-
tion of fluorine-substituted olefins,9 including elimination of
hydrogen fluoride from gem-difluorinated compounds,10 and
thermal elimination of R-fluoro-substituted sulfoxides.11
Thus, fluorine-substituted dehydroalanines might be accessed
via precursors 1 or 3 (Scheme 1). Given the strongly basic
conditions required for the elimination of hydrogen fluoride
from difluoroalanine, which could lead to racemization of
other residues in the peptide, we focused our attention on
synthon 1. R-Fluorosulfides can be prepared by the fluoro-
Pummerer rearrangement of sulfoxides with diethylamino-
sulfur trifluoride (DAST),11 by treatment of sulfides with
DAST12 or Selectfluor,13 or via the reaction of thioacetals
with mercuric fluoride.14 To evaluate these methods for the
preparation of 1, various N-protected serine derivatives were
converted to their corresponding sulfides and sulfoxides as
outlined for Fmoc-serine in Scheme 2. Fmoc-serine was
1:1 mixture of diastereomers. The direct fluorination of 6
with either Selectfluor or DAST produced 8 in lower yields
than that for the SbCl3-catalyzed15 fluoro-Pummerer rear-
rangement of the diastereomeric mixture of sulfoxides 7. The
latter reaction was carried out using the protocols developed
in the laboratories of McCarthy11 and Robins15 and produced
fluorosulfide 8 in good yield as a 1:1 mixture of diastereo-
mers. To address whether the stereochemistry at sulfur in
sulfoxide 7 had an influence on the diastereoselectivity of
the fluoro-Pummerer reaction, the two diastereomers of 7
were separated by silica gel chromatography and treated in
parallel with DAST/SbCl3 in CH2Cl2. Compound 8 was
obtained with identical diastereomer ratios for both reactions,
indicating that the stereochemistry of the reaction is not
dependent on the configuration at sulfur of the sulfoxide.
This is consistent with the proposed reaction of fluoride anion
onto a thiocarbenium intermediate formed upon reaction of
the sulfoxide with DAST.10
When this synthetic route was followed using Boc as the
N-protecting group, the reaction of sulfoxide 11 with DAST
produced 13 as a major byproduct (30%), which was not
formed in appreciable amounts with 7. Compound 13 is
presumably formed by cyclization of the carbamate carbonyl
oxygen onto the thiocarbenium intermediate. Similar cy-
clizations have been utilized intentionally in several labo-
ratories.16 A number of modifications to the reaction
conditions were explored to minimize the formation of this
for our purposes undesired byproduct. Attempts to increase
the relative rate of the intermolecular reaction with respect
to the intramolecular cyclization by increasing the fluoride
concentration via addition of CsF or tetrabutylammonium
triphenyldifluorosilicate (TBAT) resulted in inhibition of the
reaction. Increasing the overall concentration of the reaction
mixture did decrease the amount of 13 to <15 %.
Scheme 2
Fluorinated cysteine derivative 8 was oxidized with
mCPBA to yield four diastereomers of the fluorinated
(6) (a) Thornberry, N. A.; Bull, H. G.; Taub, D.; Greenlee, W. J.; Patchett,
A. A.; Cordes, E. H. J. Am. Chem. Soc. 1987, 109, 7543-7544. (b) Xu,
Y.; Abeles, R. H. Biochemistry 1993, 32, 806-11.
(7) Carrell, H. L.; Glusker, J. P.; Burger, V.; Manfre, F.; Tritsch, D.;
Biellmann, J. F. Proc. Natl. Acad. Sci. U.S.A. 1989, 86, 4440-4.
(8) Bey, P.; Fozard, J.; Lacoste, J. M.; McDonald, I. A.; Zreika, M.;
Palfreyman, M. G. J. Med. Chem. 1984, 27, 9-10.
(9) (a) Leroy, J.; Molines, H.; Wakselman, C. J. Org. Chem. 1987, 52,
290-292. (b) McCarthy, J. R.; Matthews, D. P.; Edwards, M. L.; Stemerick,
D. M.; Jarvi, E. T. Tetrahedron Lett. 1990, 31, 5449-5452. (c) Matthews,
D. P.; McCarthy, J. R. J. Org. Chem. 1990, 55, 2973-2975.
(10) Jarvi, E. T.; McCarthy, J. R.; Mehdi, S.; Matthews, D. P.; Edwards,
M. L.; Prakash, N. J.; Bowlin, T. L.; Sunkara, P. S.; Bey, P. J. Med. Chem.
1991, 34, 647-56.
(11) McCarthy, J. R.; Peet, N. P.; LeTourneau, M. E.; Inbasekaran, M.
J. Am. Chem. Soc. 1985, 107, 735-736.
(12) Robins, M. J.; Wnuk, S. F. J. Org. Chem. 1993, 58, 3800-3801.
(13) Lal, G. S. J. Org. Chem. 1993, 58, 2791-2796.
(14) Purrington, S. T.; Pittman, J. H. Tetrahedron Lett. 1987, 28, 3901-
3904.
(15) Wnuk, S. F.; Robins, M. J. J. Org. Chem. 1990, 55, 4757-4760.
(16) (a) Burrell, G.; Evans, J. M.; Jones, G. E.; Stemp, G. Tetrahedron
Lett. 1990, 25, 3649-3652. (b) Lafargue, P.; Guenot, P.; Lellouche, J. P.
Heterocycles 1995, 41, 947-958. (c) Padwa, A. Chem. Commun. 1998,
1417-1424. (d) Padwa, A.; Heidelbaugh, T. M.; Kuethe, J. T. J. Org. Chem.
1999, 64, 2038-2049. (e) Wipf, P.; Lim, S. J. Am. Chem. Soc. 1995, 117,
558-559. (f) Williams, D. R.; Brooks, D. A.; Berliner, M. A. J. Am. Chem.
Soc. 1999, 121, 4924-4925. (g) Padwa, A.; Heidelbaugh, T. M.; Kuethe,
J. T. J. Org. Chem. 2000, 65, 2368-2378. (h) Phillips, A. J.; Uto, Y.; Wipf,
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transformed into the diphenylmethyl ester 4, activated with
methylsulfonyl chloride, and treated with base to produce
dehydroalanine 5. Michael addition of 4-methoxybenzene-
thiol to 5 provided cysteine derivative 6, which was oxidized
with mCPBA to the corresponding sulfoxide 7, producing a
594
Org. Lett., Vol. 3, No. 4, 2001

sulfoxide 9.17 Subsequent thermolytic elimination in benzene
at 80 °C afforded two 3-fluorodehydroalanine isomers in 74%
yield. These two isomers were separated by silica gel
chromatography, and three approaches were used to deter-
mine the olefin geometry of the individual 3-fluorodehy-
droalanines. First, the 13C NMR spectrum of one of the
isomers, assigned to (Z)-3-fluorodehydroalanine, displayed
a doublet signal for the ester carbonyl carbon with a coupling
constant (3JC-F) of 11.6 Hz, whereas the putative (E)-isomer
showed a singlet. Similar magnetic coupling has been
reported for other fluorine-substituted olefins containing
fluorine trans to a carbonyl group.18 In addition, several
laboratories have shown for a series of 2-acylaminocrotonates
that the chemical shift of the vinyl proton cis to the acylamino
group occurs downfield relative to the vinyl proton in the
corresponding isomer.19 The relative chemical shifts of these
protons in the two isomers of 10 are consistent with our
assignment based on the carbon-fluorine coupling.20 Finally,
19F-irradiated 19F-1H NOE spectroscopy21 with (Z)-3-fluo-
rodehydroalanine showed enhancements of the signals of the
vinyl and amide protons as well as aromatic protons on the
Fmoc group, while only enhancement of the olefinic proton
was detected in (E)-3-fluorodehydroalanine.
Z-stereoisomer.22 The stereochemistry of the products was
assigned based on the chemical shift of the vinylic proton
in comparison with literature values for similar dehydro-
cysteine derivatives.23
The methodology was applied next to the synthesis of a
3-fluorodehydroalanine containing dipeptide as shown in
Scheme 4. Dipeptide 15 was synthesized in 76% yield by
Scheme 4
To evaluate the proposed mechanism of action of fluorine-
substituted dehydroalanines as presented in Scheme 1, both
isomers of 10 were reacted with p-methoxybenzenethiol in
the presence of diisopropylethylamine (Scheme 3). Clean
solution-phase techniques24 from acetyl glycine and 6.
Oxidation of the sulfide to the sulfoxide and treatment with
DAST gave the desired product 17 in 65% yield. As has
been observed in previous studies on the fluoro-Pummerer
reaction,11,12,25 a small amount (8%) of deoxygenated material
(i.e., 15) was produced in this reaction via an unknown
mechanism. Two isomers of the target 18 were obtained after
oxidation and thermal elimination at 80 °C. The E- and
Z-isomers were separated by silica gel chromatography and
proved stable at room temperature.
Scheme 3
In summary, we have developed methodology to prepare
fluorine-substituted dehydroalanines and to incorporate these
into dipeptides. Given the well-known success of fluorine-
containing pharmaceuticals,26 including fluoropeptides,27 the
current approach expands the arsenal of fluorinated moieties
conversion of Z-10 into the corresponding dehydrocysteine
derivative Z-14 was observed, consistent with an addition
elimination mechanism. The E-isomer of 10 produced
predominantly E-14, along with a small amount of the
(22) The formation of some Z-14 from E-10 may either occur during
the actual transformation or may be due to isomerization of E-14 catalyzed
by p-methoxybenzenethiol.
(23) Kakimoto, M.; Kai, M.; Kondo, K.; Hiyama, T. Chem. Lett. 1982,
527-528.
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32, 6387-6390.
(25) (a) Robins, M. J.; Wnuk, S. F. Tetrahedron Lett. 1988, 29, 5729-
5732. (b) Boys, M. L.; Collington, E. W.; Finch, H.; Swanson, S.;
Whitehead, J. F. Tetrahedron Lett. 1988, 29, 3365-3368. (c) Sufrin, J. R.;
Spiess, A. J.; Kramer, D. L.; Libby, P. R.; Porter, C. W. J. Med. Chem.
1989, 32, 997-1001. (d) Sufrin, J. R.; Spiess, A. J.; Alks, V. J. Fluor.
Chem. 1990, 49, 177-182. (e) Krishnan, G.; Sampson, P. Tetrahedron Lett.
1990, 31, 5609-5612.
(26) (a) Abeles, R. H.; Alston, T. A. J. Biol. Chem. 1990, 265, 16705-
8. (b) Welch, J. T.; Eswarakrishnan, S. Fluorine in Bioorganic Chemistry;
John Wiley & Sons, 1991. (c) Filler, R.; Kobayashi, Y.; Yagupolskii, L.
M. Organofluorine Compounds in Medicinal Chemistry and Biomedical
Applications; Elsevier: New York, 1993. (d) Ojima, I.; Welch, J. T.;
McCarthy, J. R. Biomedical Frontiers of Fluorine Chemistry; American
Chemical Society: Washington, DC, 1996.
(17) The fluorine substituted R-chiral center does not lead to significant
asymmetric induction in the oxidation. For a discussion of stereoelectronic
effects in this reaction, see Fujita, M.; Suzuki, M.; Ogata, K.; Ogura, K.
Tetrahedron Lett. 1991, 32, 1463-1466.
(18) (a) Nguyen, T.; Leroy, J.; Wakselman, C. J. Org. Chem. 1993, 58,
3772-3774. (b) Miura, K.; Sugimoto, J.; Oshima, K. Bull. Chem. Soc. Jpn.
1992, 65, 1513-1521. (c) Boguslavskaya, L. S.; Chernov, A. N.; Krom, E.
N. J. Org. Chem. USSR 1980, 1392-1397.
(19) (a) Brown, A. G.; Smale, T. C. J. Chem. Soc. D 1969, 1489-1490.
(b) Srinivasan, A.; Richards, K. D.; Olsen, R. K. Tetrahedron Lett. 1976,
18, 891-894. (c) Effenberger, F.; Ku¨hlwein, J.; Drauz, K. Liebigs Ann.
Chem. 1993, 1295-1301.
(20) The chemical shifts of the vinyl protons of (E)-3-fluorodehydro-
alanine and (Z)-3-fluorodehydroalanine are 8.18 ppm (2JHF ) 78.1 Hz) and
7.66 ppm (2JHF ) 72.6 Hz), respectively.
(21) McCarthy, J. R.; Huber, E. W.; Le, T.-B.; Laskovics, F. M.;
Matthews, D. P. Tetrahedron 1996, 52, 45-58.
(27) Imperiali, B. AdV. Biotechnol. Processes 1988, 10, 97-129.
Org. Lett., Vol. 3, No. 4, 2001
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that can be used in the preparation of bioactive molecules.
Efforts to introduce these reactive Michael acceptors into
natural products are underway and will be reported in due
course.
Center for Excellence, funded in part by the W. M. Keck
Foundation, NIH (PHS 1 S10 RR10444), and NSF (CHE
96-10502), and Prof. Peter Petillo for helpful discussions
regarding the stereochemical assignments of 10.
Supporting Information Available: Experimental pro-
cedures for all transformations that produced previously
unknown compounds as well as their full spectral charac-
terization. This material is available free of charge via the
Internet at http://pubs.acs.org.
Acknowledgment. This work was supported by grants
from the Burroughs-Wellcome Fund (APP 1920) and the
National Institutes of Health (GM58822). W.A.V. thanks 3M
for a nontenured faculty grant. We thank Drs. Vera Mainz
and Paul Molitor for assistance with the heteronuclear NOE
experiments, obtained in the Varian Oxford Instrument
OL006997S
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Org. Lett., Vol. 3, No. 4, 2001