Synthesis and 12-helical secondary structure of β-peptides containing (2R,3R)-Aminoproline

Article abstract of DOI:10.1021/ol0266370

equation presented (2R,3R)-Aminoproline, a pyrrolidine-based β-amino acid, was synthesized and incorporated into hexa-β-peptide 4. This residue confers water solubility when the ring nitrogen is protonated and allows for 12-helix formation in aqueous solu

Full text of DOI:10.1021/ol0266370

ORGANIC
LETTERS
2002
Vol. 4, No. 19
3317-3319
Synthesis and 12-Helical Secondary
Structure of â-Peptides Containing
(2R,3R)-Aminoproline
Emilie A. Porter, Xifang Wang, Margaret A. Schmitt, and Samuel H. Gellman*
Department of Chemistry, UniVersity of Wisconsin, Madison, Wisconsin 53706
gellman@chem.wisc.edu
Received July 30, 2002
ABSTRACT
(2R,3R)-Aminoproline, a pyrrolidine-based â-amino acid, was synthesized and incorporated into hexa-â-peptide 4. This residue confers water
solubility when the ring nitrogen is protonated and allows for 12-helix formation in aqueous solution. Circular dichroism spectra display the
12-helical signature, and 12-helical structure was confirmed by 2D NMR analysis.
Oligomers that are capable of taking on well-defined
conformations in solution (“foldamers”) have received much
attention in recent years.1 One class of foldamers, â-peptides,
has been studied by several research groups.2 Recently,
â-peptides have been found to display useful biological
functions.3 Some of the biologically active â-peptides that
have come from our laboratory display a 12-helical secondary
structure. The 12-helix is promoted by â-amino acids that
are constrained by five-membered rings, such as trans-
aminocyclopentanecarboxylic acid (ACPC, Figure 1).4,5 This
(1) (a) Gellman, S. H. Acc. Chem. Res. 1998, 31, 173-180. (b) Barron,
A. E.; Zuckermann, R. N. Curr. Opin. Chem. Biol. 1999, 3. (c) Hill, D. J.;
Mio, M. J.; Prince, R. B.; Hughes, T. S.; Moore, J. S. Chem. ReV. 2001,
101, 3893-4011.
(2) (a) Cheng, R. P.; Gellman, S. H.; DeGrado, W. F. Chem. ReV. 2001,
101, 3219-3232. (b) Gademann, K.; Hintermann, T.; Schreiber, J. V. Curr.
Med. Chem. 1999, 6, 905-925. (c) Claridge, T. D. W.; Goodman, J. M.;
Moreno, A.; Angus, D.; Barker, S. F.; Taillefumier, C.; Watterson, M. P.;
Fleet, G. W. J. Tetrahedron Lett. 2001, 42, 4251-4255.
Figure 1. Protected monomers for 12-helical â-peptides.
helix is defined by 12-membered ring hydrogen bonds [Cd
O(i) f N-H(i + 3)] and has approximately 2.6 residues
per turn. The 12-helix is well-suited for biological applica-
(3) (a) Werder, M.; Hauser, H.; Abele, S.; Seebach, D. HelV. Chim. Acta
1999, 82, 1774-1783. (b) Gademann, K.; Ernst, M.; Hoyer, D.; Seebach,
D. Angew. Chem., Int. Ed. 1999, 38, 1223-1226. (c) Hamuro, Y.; Schneider,
J. P.; DeGrado, W. F. J. Am. Chem. Soc. 1999, 121, 12200-12201. (d)
Porter, E. A.; Wang, X.; Lee, H. S.; Weisblum, B.; Gellman, S. H. Nature
2000, 404, 565. (e) Gademann, K.; Ernst, M.; Seebach, D.; Hoyer, D. HelV.
Chim. Acta 2000, 83, 16-33. (f) Gademann, K.; Seebach, D. HelV. Chim.
Acta 2001, 84, 2924-2937. (g) Gademann, K.; Kimmerlin, T.; Hoyer, D.;
Seebach, D. J. Med. Chem. 2001, 44, 2460-2468. (h) Liu, D. H.; DeGrado,
W. F. J. Am. Chem. Soc. 2001, 123, 7553-7559. (i) Arvidsson, P. I.;
Frackenpohl, J.; Ryder, N. S.; Liechty, B.; Petersen, F.; Zimmermann, H.;
Camenisch, G. P.; Woessner, R.; Seebach, D. ChemBioChem 2001, 2, 771.
(j) Umezawa, N.; Gelman, M. A.; Haigis, M. C.; Raines, R. T.; Gellman,
S. H. J. Am. Chem. Soc. 2002, 124, 368-369. (k) LePlae, P. R.; Fisk, J.
D.; Porter, E. A.; Weisblum, B.; Gellman, S. H. J. Am. Chem. Soc. 2002,
124, 6820-6821. (l) Rueping, M.; Mahajan, Y.; Sauer, M.; Seebach, D.
ChemBioChem 2002, 3, 257-259. (m) Porter, E. A.; Weisblum, B.;
Gellman, S. H. J. Am. Chem. Soc. 2002, 124, 7324-7330. (n) Arnold, U.;
Hinderaker, M. P.; Nilsson, B. L.; Huck, B. R.; Gellman, S. H.; Raines, R.
T. J. Am. Chem. Soc. 2002, 124, 8522-8523.
(4) (a) Appella, D. H.; Christianson, L. A.; Klein, D. A.; Powell, D. R.;
Huang, X. L.; Barchi, J. J.; Gellman, S. H. Nature 1997, 387, 381-384.
(b) Appella, D. H.; Christianson, L. A.; Klein, D. A.; Richards, M. R.;
Powell, D. R.; Gellman, S. H. J. Am. Chem. Soc. 1999, 121, 7574-7581.
(c) Another type of constrained â-amino acid has been reported to promote
the 12-helix: Winkler, J. D.; Piatnitski, E. L.; Mehlmann, J.; Kasparec, J.;
Axelsen, P. H. Angew. Chem., Int. Ed. 2001, 40, 743-745.
10.1021/ol0266370 CCC: $22.00 © 2002 American Chemical Society
Published on Web 08/29/2002

tions because its dimensions are similar to those of the
R-helix found in proteins.
for ACPC by Tilley et al.9 The stereocenters are set by a
nonfermenting baker’s yeast reduction10 of known ketoester
2;11 the reaction proceeds in approximately 89% ee (deter-
mined by chiral HPLC) and excellent yield. However, as a
result of tedious workup, the yield tends to decrease upon
reaction scale-up. The absolute configuration of the hydroxy-
ester product has been established by Cooper et al.11b The
alcohol can be transformed to an azide by a Mitsunobu
reaction with hydrazoic acid in excellent yield. Gomez-Vidal
and Silverman have reported using DPPA in the synthesis
of the methyl ester version of the molecule.12 While DPPA
is a more convenient reagent than hydrazoic acid, in this
case the yields are only moderate (60-75%), and an
elimination byproduct inseparable from the desired azide
(∼10%) is formed during the DPPA reaction. The azide is
reduced to an amine with tin(II) chloride and protected with
a carboxybenzyl group. The ester is hydrolyzed and the
carboxybenzyl group is replaced by a fluorenylmethyoxy-
carbonyl group for solid-phase synthesis.
We have previously shown that 12-helices can form in
water when a pyrrolidine monomer, trans-3-aminopyrroli-
dine-4-carboxylic acid (APC, Figure 1), is incorporated.6
APC confers water solubility by virtue of protonation of the
ring nitrogen. To have maximum flexibility in the design of
â-peptides for biological applications, we need a pool of
constrained â-amino acids with variety in the type or
orientation of peripheral functional groups. Here we describe
the synthesis of an isomer of APC, (2R,3R)-aminoproline
(AP), which differs from APC in the position of the
pyrrolidine nitrogen relative to the substituents but neverthe-
less promotes 12-helix formation.
Our synthesis of enantiomerically pure AP in a protected
form is outlined in Scheme 1. We were not able to synthesize
Scheme 1. Synthesis of Fmoc-AP(Boc) 1
Compound 1 was used along with Fmoc-ACPC and Fmoc-
APC(Boc) in standard automated solid-phase peptide syn-
thesis of several â-peptide hexamers, including 3 and 4
(Figure 2), with HBTU activation. Treatment with 95% TFA
Fmoc-AP(Boc) 1 by a route analogous to those used for
Fmoc-APC(Boc)7 and Fmoc-ACPC;8 the starting ketoester
2 would not react with methylbenzylamine to form the
enamine necessary for the key reductive amination, even
under Dean-Stark conditions (Scheme 2). The synthesis we
employed for 1 is analogous to a route previously described
Figure 2. â-peptides 3 and 4. Curved arrows superimposed on 4
indicate NOEs between residues that are not adjacent in sequence
(CD3OH). The dashed arrow indicates an NOE that is ambiguous
because of resonance overlap.
cleaved the â-peptide from the resin and deprotected the
pyrrolidine nitrogens. Although the baker’s yeast reduction
Scheme 2
(5) cis-ACPC promotes strand formation: Martinek, T. A.; To´th, G. K.;
Vass, E.; Hollo´si, M.; Fu¨lo¨p, F. Angew. Chem., Int. Ed. 2002, 41, 1718-
1721.
(6) Wang, X.; Espinosa, J. F.; Gellman, S. H. J. Am. Chem. Soc. 2000,
122, 4821-4822.
(7) Lee, H. S.; LePlae, P. R.; Porter, E. A.; Gellman, S. H. J. Org. Chem.
2001, 66, 3597-3599.
(8) LePlae, P. R.; Umezawa, N.; Lee, H. S.; Gellman, S. H. J. Org. Chem.
2001, 66, 5629-5632.
(9) Tilley, J. W.; Danho, W.; Shiuey, S.; Kulesha, I.; Swistok, J.;
Makofske, R.; Michalewsky, J.; Triscari, J.; Nelson, D.; Weatherford, S.;
Madison, V.; Fry, D.; Cook, C. J. Med. Chem. 1992, 35, 3774.
(10) Seebach, D.; Roggo, S.; Maetzke, T.; Braunschweiger, H.; Cercus,
J.; Krieger, M. HelV. Chim. Acta 1987, 70, 1605-1615.
(11) (a) Blake, J.; Willson, C. D.; Rapoport, H. J. Am. Chem. Soc. 1964,
86, 5293-5299. (b) Cooper, J.; Gallagher, P. T.; Knight, D. W. J. Chem.
Soc., Perkin Trans. 1 1993, 1313-1317.
3318
Org. Lett., Vol. 4, No. 19, 2002

in the preparation of 1 proceeded with only 89% ee, the
â-peptides described here are diastereomerically pure after
HPLC purification. Initial examination of hexamer 3 in
222 nm in methanol, and the maximum is blue-shifted to
201 nm in aqueous solution. The CD spectrum in methanol
is more intense, suggesting a higher population of 12-helix
in methanol than in aqueous solution. Helix stabilization by
alcohols relative to water has been reported previously both
for â-peptides2a and conventional R-peptides.14 It is signifi-
cant that hexamer 4 forms a 12-helix in aqueous solution,
because R-peptides with six amino acids, as well as most
short â-peptides comprised of acyclic residues,15 do not form
helices in water. We have previously shown that â-peptides
consisting of ACPC, APC and related residues form the 12-
helix in water with only six residues.4,16
The AP residue reported here adds to our monomer pool
for the synthesis of functionalized 12-helical â-peptides. AP
has recently been incorporated into a 17-residue â-peptide
that displays antimicrobial properties.3m We anticipate that
the AP residue will be valuable in the development of
additional â-peptides with useful properties.
1
CD3OH via NMR revealed extensive overlap among H
resonances, which precluded further analysis. Hexamer 4 was
designed to show enhanced 1H resonance dispersion; 4
contains three different types of â-amino acid residue, while
3 contains only two. Two-dimensional NMR (ROESY) data
for 4 in CD3OH allowed us to identify eight NOEs between
residues that are not adjacent in the sequence, all of which
are consistent with at least partial population of the 12-helical
conformation (no NOE inconsistent with the 12-helix was
detected). Two types of nonsequential NOEs were observed,
CâHi f NHi+2 (four of five detected; one is ambiguous) and
CâHi f CRHi+2 (all four detected). These results show that
the AP residue promotes 12-helix formation in solution, since
many of the NOEs involve or span the AP residues.
â-Peptide hexamers 3 and 4 were also characterized by
circular dichroism for comparison with APC/ACPC â-pep-
tides4 and all-ACPC â-peptides.7 Hexamer 4 showed the
characteristic CD signature for the 12-helix in both methanol
and aqueous buffer (10 mM Tris, pH 7.2, Figure 3).13 Very
similar data were obtained for 3 (not shown). The CD
spectrum of 4 has a maximum at 204 nm and a minimum at
Acknowledgment. This research was supported by the
NIH (GM56414). E.A.P. was supported in part by a
Biotechnology Training Grant (NIH 5 T32 GM08349).
M.A.S. was supported in part by a Biophysics Training
Grant. CD data were obtained in the Biophysics Instrumenta-
tion Facility at UW-Madison, supported by NSF. NMR
spectra were obtained in the Chemistry Instrument Center,
supported by NSF and NIH.
Supporting Information Available: Experimental pro-
cedure for compound 1, oligomer synthesis, and 2D NMR
data. This material is available free of charge via the Internet
at http://pubs.acs.org.
OL0266370
(12) Gomez-Vidal, J. A.; Silverman, R. B. Org. Lett. 2001, 3, 2481-
2484.
(13) Applequist, J.; Bode, K. A.; Appella, D. H.; Christianson, L. A.;
Gellman, S. H. J. Am. Chem. Soc. 1998, 120, 4891-4892.
(14) Creighton, T. E. Proteins: Structures and Molecular Properties,
2nd ed.; W. H. Freeman and Company: New York, 1993.
(15) A few â-peptides containing exclusively acyclic residues have been
reported to be 14-helical in aqueous solution. (a) A seven-residue â-pep-
tide: Gung, B. W.; Zou, D.; Stalcup, A. M.; Cottrell, C. E. J. Org. Chem.
1999, 64, 2176-2177. (b) A seven-residue â-peptide stabilized by salt
bridges: Arvidsson, P. I.; Rueping, M.; Seebach, D. Chem. Commun. 2001,
649-650. (c) A 15-residue â-peptide stabilized by salt bridges: Cheng, R.
P.; DeGrado, W. F. J. Am. Chem. Soc. 2001, 123, 5162-5163.
(16) Lee, H. S.; Syud, F. A.; Wang, X.; Gellman, S. H. J. Am. Chem.
Soc. 2001, 123, 7721-7722.
Figure 3. CD spectra of 4 (0.2 mM) in methanol and in aqueous
buffer.
Org. Lett., Vol. 4, No. 19, 2002
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