12-helix folding of cyclobutane β-amino acid oligomers

Article abstract of DOI:10.1021/ol101267u

The hexamer and octamer of trans-2-aminocyclobutane carboxylic acid were prepared and their conformational preferences studied experimentally and using molecular modeling. All observations suggest a marked preference for the folding of these oligomers int

Full text of DOI:10.1021/ol101267u

ORGANIC
LETTERS
2010
Vol. 12, No. 16
3606-3609
12-Helix Folding of Cyclobutane
ꢀ-Amino Acid Oligomers
Carlos Fernandes,† Sophie Faure,† Elisabeth Pereira,† Vincent The´ry,†
Vale´rie Declerck,‡ Re´gis Guillot,‡ and David J. Aitken*,‡
Clermont UniVersite´, UniVersite´ Blaise Pascal, Laboratoire SEESIB (UMR 6504 - CNRS),
24 aVenue des Landais, 63177 Aubie`re cedex, France, and UniVersite´ Paris-Sud 11,
Laboratoire de Synthe`se Organique & Me´thodologie, ICMMO (UMR 8182 - CNRS),
15 rue Georges Clemenceau, 91405 Orsay cedex, France
daVid.aitken@u-psud.fr
Received June 2, 2010
ABSTRACT
The hexamer and octamer of trans-2-aminocyclobutane carboxylic acid were prepared and their conformational preferences studied experimentally
and using molecular modeling. All observations suggest a marked preference for the folding of these oligomers into a well-defined 12-helical
conformation, in both solution and the solid state.
There is considerable interest in synthetic oligomers which
have well-defined folding preferences (foldamers).1 ꢀ-Pep-
tides are among the most studied class of compounds in this
respect and offer an array of possibilities for regular folding
through hydrogen bond formation.2 Some ground rules have
emerged for the folding of oligomers which contain ꢀ-amino
acids (ꢀ-AAs): the 14-helix is a preferred conformation in
oligomers of ꢀ-AAs with a six-membered ring constraint,
such as trans-2-aminocyclohexane carboxylic acid,3 while
oligomers of trans-2-aminocyclopentane carboxylic acid
(trans-ACPC) and related five-membered heterocyclic ꢀ-AAs
have distinct 12-helix preferences.4 Mixed R/ꢀ-peptides
incorporating carbocyclic ꢀ-AAs display other conforma-
tional preferences.5
† Clermont Universite´.
‡ Universite´ Paris-Sud 11.
(1) (a) Hecht, S.; Huc, I.; Foldamers: Structure, Properties, and
Applications; Wiley-VCH: Weinheim, 2007. (b) Seebach, D.; Gardiner, J.
Acc. Chem. Res. 2008, 41, 1366. (c) Hill, D. J.; Mio, M. J.; Prince, R. B.;
Hughes, T. S.; Moore, J. S. Chem. ReV. 2001, 101, 3893. (d) Gellman,
S. H. Acc. Chem. Res. 1998, 31, 173. (e) Li, X.; Yang, D. Chem. Commun.
2006, 585. (f) Gong, B. Acc. Chem. Res. 2008, 41, 1376. (g) Li, Z. T.;
Hou, J. L.; Li, C.; Yi, H. P. Chem. Asian J. 2006, 1, 766.
The folding propensities of oligomers constructed from
four-membered cyclic ꢀ-AA building blocks are less clear-
(3) (a) Appella, D. H.; Christianson, L. A.; Karle, I. L.; Powell, D. R.;
Gellman, S. H. J. Am. Chem. Soc. 1999, 121, 6206. (b) Schinnerl, M.;
Murray, J. K.; Langenhan, J. M.; Gellman, S. H. Eur. J. Org. Chem. 2003,
721. (c) Hete´nyi, A.; Ma´ndity, I. M.; Martinek, T. A.; To´th, G. K.; Fu¨lo¨p,
F. J. Am. Chem. Soc. 2005, 127, 547.
(2) (a) Seebach, D.; Abele, S.; Gademann, K.; Guichard, G.; Hintermann,
T.; Jaun, B.; Matthews, J. L.; Schreiber, J. V.; Oberer, L.; Hommel, U.;
Widmer, H. HelV. Chim. Acta 1998, 81, 932. (b) Martinek, T. A.; Ma´ndity,
I. M.; Fu¨lo¨p, L.; To´th, G. K.; Vass, E.; Hollo´si, M.; Forro´, E.; Fu¨lo¨p, F.
J. Am. Chem. Soc. 2006, 128, 13539. (c) Threlfall, R.; Davies, A.; Howarth,
N. M.; Fisher, J.; Cosstick, R. Chem. Commun. 2008, 585. (d) Kritzer, J. A.;
Hodsdon, M. E.; Schepartz, A. J. Am. Chem. Soc. 2005, 127, 4118. (e)
Seebach, D.; Beck, A. K.; Bierbaum, D. J. Chem. BiodiVersity 2004, 1,
1111. (f) Chakraborty, P.; Diederichsen, U. Chem.sEur. J. 2005, 11, 3207.
(g) Cheng, R. P.; Gellman, S. H.; DeGrado, W. F. Chem. ReV. 2001, 101,
3893.
(4) (a) Lee, H.-S.; Syud, F. A.; Wang, X.; Gellman, S. H. J. Am. Chem.
Soc. 2001, 123, 7721. (b) Appella, D. H.; Christianson, L. A.; Klein, D. A.;
Powell, D. R.; Huang, X.; Barchi, J. J.; Gellman, S. H. Nature 1997, 387,
381. (c) 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. (d)
Wang, X.; Espinosa, J. F.; Gellman, S. H. J. Am. Chem. Soc. 2000, 122,
4821.
10.1021/ol101267u  2010 American Chemical Society
Published on Web 07/21/2010

cut. Fleet6 described an oxetane-based cis-ꢀ-AA hexamer
which adopted a 10-helix conformation, while Ortun˜o
observed 14-helical folding for a tetramer containing in
alternation two cis-cyclobutane ꢀ-AA (cis-ACBC) residues
and two ꢀ-alanines.7 Homo-oligomers composed uniquely
of cis-ACBC units (2-8 residues) showed a conformational
bias derived essentially from six-membered intramolecular
hydrogen bonds, which facilitated supramolecular interactions
and self-assembly, producing nanosized fibers or gels.8 How-
ever, trans-ACBC units have only been studied in dipeptides:
a preference for the formation of eight-membered hydrogen-
bond rings was noted for (trans,cis) and (trans,trans) combina-
tions,9 provoking the question of whether this structural feature
might prevail in longer oligomers of trans-ACBC. It thus
seemed opportune to examine the conformational behavior of
such ꢀ-peptides.
Figure 2. Circular dichroism spectra in MeOH (0.2 mM) for dimer
1, tetramer 2, hexamer 3, and octamer 4. Molar ellipticity has been
The preparation of the target oligomers was not a trivial
task since the parent ꢀ-AA is only moderately stable10 and
has only recently been described in enantiomerically pure
form.11 However, solution-phase oligomer assembly was
achieved successfully and is summarized as follows. Dimer
1 was obtained by coupling equimolar amounts of the N-Boc
and the methyl ester derivatives of (1R,2R)-trans-ACBC.
This is the only step requiring any particular care: the free
amine of the methyl ester of (1R,2R)-trans-ACBC is liberated
from its stable hydrochloride immediately prior to coupling.
Subsequent chain extension procedures were free of dif-
ficulty: selective deprotection and coupling of two dimer units
gave the tetramer 2. Analogous operations were performed
to combine 1 and 2 to provide hexamer 3 and to combine 2
equiv of 2 to give octamer 4 (full details are given in the
Supporting Information file). In contrast to the free monomer,
all of these terminally protected oligomers are stable, and
their structures are shown in Figure 1.
normalized for concentration and per-residue.
1
which allowed complete assignment of all backbone H
signals, a necessary prerequisite for the intended NOE
experiments. The two N-terminal amide and carbamate
protons were deshielded, which can be explained as a result
of these hydrogens being less involved in intramolecular
CdO···H-N hydrogen bonding, leaving them more solvent-
exposed and forming hydrogen bonds with the solvent. The
temperature coefficients for the 1H NMR amide (or carbam-
ate) signals of oligomers 3 and 4 were consistent with this
postulate: in each case, the values for the protons of residues
1 and 2 (-15 and -12 ppb/°C, respectively) were consider-
ably larger than all others (<-6 ppb/°C).
ROESY spectral data were then used to identify nonad-
jacent NOE interactions, and the three correlation types
observed in both 3 and 4 (CꢀHi-NHi+2, CꢀHi-CRHi+2, CꢀHi-
NHi+3) are characteristic of a 12-helix.4 Correlations are
summarized in Figure 3 for hexamer 3.
NMR data obtained for 3 were used in molecular modeling
studies. Molecular dynamics calculations were carried out
(5) (a) Horne, W. S.; Gellman, S. H. Acc. Chem. Res. 2008, 41, 1399.
(b) Baldauf, C.; Gunther, R.; Hofmann, H. J. Biopolymers 2006, 84, 408.
(c) Ma´ndity, I. M.; We´ber, E.; Martinek, T. A.; Olajos, G.; To´th, G. K.;
Vass, E.; Fu¨lo¨p, F. Angew. Chem., Int. Ed. 2009, 48, 2171. (d) De Pol, S.;
Zorn, C.; Klein, C. D.; Zerbe, O.; Reiser, O. Angew. Chem., Int. Ed. 2004,
43, 511.
Figure 1. Parent trans-ꢀ-amino acid building block and the series
of oligomers.
(6) 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.
Circular dichroism data gave the first indication of a
change in conformational preferences with increasing oli-
gomer length (Figure 2). In MeOH solution, a marked
evolution in behavior was observed progressing from dimer
1 through tetramer 2 to hexamer 3. The spectra of hexamer
3 and octamer 4 were very similar, with a maximum near
209 nm and a minimum near 227 nm. While it has been
pointed out that CD data on its own are insufficient for
drawing firm conclusions on the conformational preferences
of ꢀ-peptides,12 we noted that the data for 3 and 4 resembled
a slightly red-shifted version of the characteristic spectral
signature of a 12-helix in the same solvent.4
(7) Izquierdo, S.; Kogan, M. J.; Parella, T.; Moglioni, A. G.; Branchadell,
V.; Giralt, E.; Ortun˜o, R. M. J. Org. Chem. 2004, 69, 5093.
(8) (a) Torres, E.; Gorrea, E.; Burusco, K. K.; Da Silva, E.; Nolis, P.;
Ru´a, F.; Boussert, S.; D´ıez-Pe´rez, I.; Dannenberg, S.; Izquierdo, S.; Giralt,
E.; Jaime, C.; Branchadell, V.; Ortun˜o, R. M. Org. Biomol. Chem. 2010, 8,
564. (b) Ru´a, F.; Boussert, S.; Parella, T.; D´ıez-Pe´rez, I.; Branchadell, V.;
Giralt, E.; Ortun˜o, R. M. Org. Lett. 2007, 9, 3643.
(9) (a) Torres, E.; Gorrea, E.; Da Silva, E.; Nolis, P.; Branchadell, V.;
Ortun˜o, R. M. Org. Lett. 2009, 11, 2301. (b) Izquierdo, S.; Ru´a, F.; Sbai,
´
´
A.; Parella, T.; Alvarez-Larena, A.; Branchadell, V.; Ortun˜o, R. M. J. Org.
Chem. 2005, 70, 7963.
(10) trans-ACBC is prone to ring opening in solution, similarly to cis-
ACBC: Aitken, D. J.; Gauzy, C.; Pereira, E. Tetrahedron Lett. 2004, 45,
2359.
(11) (a) Fernandes, C.; Pereira, E.; Faure, S.; Aitken, D. J. J. Org. Chem.
2009, 74, 3217. (b) Fernandes, C.; Gauzy, C.; Yang, Y.; Roy, O.; Pereira,
E.; Faure, S.; Aitken, D. J. Synthesis 2007, 2222.
The best solubility and NMR spectral resolution of 3 and
4 were secured in pyridine-d5, which was the only solvent
(12) Daura, X.; Bakowies, D.; Seebach, D.; Fleischhauer, J.; Van
Gunsteren, W. F.; Kru¨ger, P. Eur. Biophys. J. 2003, 32, 661.
Org. Lett., Vol. 12, No. 16, 2010
3607

5). Six standard hydrogen bonds form the basis of the helical
structure, with CdO···H-N distances within the range
2.03-2.22 Å and O···H-N angles falling in the range
133.9-160.1°. The cyclobutane ring conformation is highly
uniform all along the oligomer chain, with a puckering angle
in the range 25.6-29.4°, with exceptions at residue 5 and
8, where puckering of 33.2° and 31.8°, respectively, are
observed. The amine and carbonyl moieties occupy pseu-
doequatorial positions. Figure 5 shows a typical constituent
trans-ACBC moiety (residue 3).
Figure 3. Graphical summary of nOes observed for hexamer 3.
Four molecules are present in the asymmetric unit of the
crystal, and their conformations are very similar. Two
molecules are aligned end-to-end on the same axis, but no
hydrogen bond exists between them; the other two molecules
are also in an antiparallel orientation but on a different axis.
Intermolecular hydrogen bonds are formed between helices
which are on different axes but having the same global
orientation. In addition to the six intramolecular hydrogen
bonds that maintain the helix, each helix forms four
intermolecular hydrogen bonds with the closest neighbors;
all hydrogen-bonding sites are thus satisfied. The second pair
of helices in the asymmetric unit pack via the same set of
intermolecular hydrogen bonds but are in an opposite global
orientation, thus creating layers of helices; there is no
apparent interaction between these layers. Likewise, there
is little side-to-side intermolecular contact between the
cyclobutane rings in neighboring layers.
Analogous correlations were observed for octamer 4. Conditions:
10 mM solution in pyridine-d5, 298 K, mixing time 300 ms.
on hexamer 3 in a pyridine continuum using SYBYL 7.0
software and the MMFFs94 force field, initially without
restraints. Five low-energy conformer families emerged, and
each was subjected to an ab initio geometrical optimization
using the B3LYP STO-3G base. This refinement provided
only two accessible conformer families, from which the
lowest potential energy corresponded to a 12-helix.13 Re-
strained molecular dynamics calculations were then carried
out in a similar manner, including energy terms for the
ROESY distance restraints and dihedral angle restraints based
on NHi-CꢀHi coupling constants. Only 12-helical structures
were obtained; the minimum energy conformer is shown in
Figure 4.
It is interesting to compare the 12-helices adopted by the
trans-ACBC oligomers in the crystalline state and the
modeled solution state. Figure 6 shows the superposition of
two comparable internal segments from the minimum energy
conformer for 3 and the crystal structure for 4. Overall, there
is a good overlap of the structural features, including the
conformation of the cyclobutane rings. In the solid state, the
backbone torsion angles are marginally larger, leading to a
slight elongation of the 12-helix.
The solid state conformation of the backbone of molecule
4 is very similar to that of a protected octamer of trans-
ACPC,4c while more subtle differences appear in the behavior
of the peripheral parts of the respective carbocycles. There
is some lateral interaction between cyclopentane rings of
neighboring trans-ACPC molecules, whereas the rigidity of
the cyclobutane rings in 4 appears to preclude such interac-
tions.
In summary, the hexamer and octamer of trans-ACBC do
not fold around short-range (eight-membered ring) hydrogen
bonds but show instead a clear preference for longer-range
hydrogen bonding interactions leading to a stabilized 12-
helix. This structure is an important manifold for the
elaboration of functional foldamers: antibiotic and antiviral
activities14 and γ-secretase inhibition15 have been reported
for rationally designed 12-helix oligomers, and efforts are
made to secure new ways of stabilizing this conformational
Figure 4. Minimum energy structure for hexamer 3 showing the
hydrogen bonds implicated in the 12-helix structure (top) and
looking down the helix axis (bottom).
Crystals of octamer 4 grown from CHCl3/Et2O solution
were suitable for X-ray diffraction. A well-defined 12-helical
structure is adopted by 4 in the solid state (Figure 5). The
average backbone torsion angle is -95.7°, with little
distortion (max. -100.1° at residue 4, min. -89.7° at residue
(14) (a) Porter, E. A.; Wang, X.; Lee, H.-S.; Weisblum, B.; Gellman,
S. H. Nature 2000, 404, 565. (b) English, E. P.; Chumanov, R. S.; Gellman,
S. H.; Compton, T. J. Biol. Chem. 2006, 281, 2661.
(13) In response to a request by a reviewer, the lowest-energy conformer
was reoptimized at the 6-31G(d,p) level, with no discernable difference in
the core helical structure. See the Supporting Information for details.
(15) Imamura, Y.; Watanabe, N.; Umezawa, N.; Iwatsubo, T.; Kato,
N.; Tomita, T.; Higuchi, T. J. Am. Chem. Soc. 2009, 131, 7353.
3608
Org. Lett., Vol. 12, No. 16, 2010

Figure 5. Crystal structure images of octamer 4. For clarity, all hydrogen atoms have been removed except those involved in hydrogen bonds.
Top left: a single molecule is shown with the six H-bonds which stabilize the 12-helix. Top right: an enlargement of one of the ꢀ-amino acid units
(residue 3), highlighting the cyclobutane ring conformation. Bottom: Intermolecular interactions of a selected molecule with four neighbors: the
NHs of residues 1 and 2 interact with the COs of residues 6 and 7 (respectively) of the two closest helices in adjacent planes; likewise, the COs
of residues 6 and 7 interact with the NHs of residues 1 and 2 of the two nearest helices, also in adjacent planes.
feature.16 trans-ACBC-based peptides are similar, but not
identical, to trans-ACPC oligomers and thus enlarge the set
of basic structures available for foldamer development and
design.
Acknowledgment. This work was supported by a graduate
research grant (to C.F.) from the French Ministry of
Research. We thank Dr. C. T. Craescu (Institut Curie, Orsay)
for help with CD.
Supporting Information Available: Preparative proce-
dures, copies of NMR spectra, chemical shift assignments,
details of ROESY experiments, molecular modeling, and
crystallographic data. This material is available free of charge
via the Internet at http://pubs.acs.org.
Figure 6. Superposition of comparable segments of modeled
solution state (multicolored) and solid state (yellow) 12-helix
OL101267U
structures. The solution state segment is residues 2-5 of hexamer
3, and the solid state segment is residues 3-6 of octamer 4.
´
(16) Hete´nyi, A.; Szakonyi, Z.; Ma´ndity, I. M.; Szolnoki, E.; To´th, G. K.;
Martinek, T. A.; Fu¨lo¨p, F. Chem. Commun. 2009, 177.
Org. Lett., Vol. 12, No. 16, 2010
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