Oligomers of a 5-carboxy-methanopyrrolidine β-amino acid. A search for order

Article abstract of DOI:10.1021/ol1022917

CD spectra for homooligomers (n = 4, 6, 8) of (1S,4R,5R)-5-syn-carboxy-2- azabicyclo[2.1.1]hexane (MPCA), a methano-bridged pyrrolidine β-carboxylic acid, suggest an ordered secondary structure. Even in the absence of internal hydrogen bonding, solution N

Full text of DOI:10.1021/ol1022917

ORGANIC
LETTERS
2010
Vol. 12, No. 23
5438-5441
Oligomers of a
5-Carboxy-methanopyrrolidine ꢀ-Amino
Acid. A Search for Order
Grant R. Krow,*,† Nian Liu,† Matthew Sender,† Guoliang Lin,† Ryan Centafont,†
Philip E. Sonnet,† Charles DeBrosse,† Charles W. Ross III,‡ Patrick J. Carroll,§
Matthew D. Shoulders,| and Ronald T. Raines⊥
Department of Chemistry, Temple UniVersity, Philadelphia, PennsylVania 19122,
United States, Department of Medicinal Chemistry, Merck Research Laboratories,
Merck & Co., Inc., West Point, PennsylVania 19486-004, United States, Department of
Chemistry, UniVersity of PennsylVania, Philadelphia, PennsylVania 19104, United
States, Departments of Chemistry and Biochemistry, UniVersity of Wisconsin-Madison,
Madison, Wisconsin 53706, United States
grkrow@temple.edu
Received September 23, 2010
ABSTRACT
CD spectra for homooligomers (n ) 4, 6, 8) of (1S,4R,5R)-5-syn-carboxy-2-azabicyclo[2.1.1]hexane (MPCA), a methano-bridged pyrrolidine
ꢀ-carboxylic acid, suggest an ordered secondary structure. Even in the absence of internal hydrogen bonding, solution NMR, X-ray, and in
silico analyses of the tetramer are indicative of conformations with trans-amides and C5-amide-carbonyls oriented toward the C4 bridgehead.
This highly constrained ꢀ-amino acid could prove useful in the ongoing development of well-defined foldamers.
Among naturally occurring R-amino acids, proline 1 is the
only secondary amine. Although this means that homooli-
gomers of proline do not have N-H protons available to
stabilize secondary structures by a hydrogen bond, polypro-
line still forms ordered oligomers.1 In the course of the
rational design of ꢀ-peptides related to natural peptides,
Seebach2 wondered if ꢀ-peptidic chains without backbone
H-bonds might also fold into stable secondary structures. To
explore this possibility, Seebach and co-workers prepared
two methylene homologues of proline by formal insertion
of a methylene either between the carbonyl C-atom and the
C(R)-atom to give 2 or between the C(R)- and C(ꢀ)-atoms
to give 3. Oligomers (n ) 3, 6, 12, 18) of (R)-2 with terminal
N-Boc and OBn groups all gave CD patterns in methanol
with an intense minimum at 202 nm and a maximum at 223
that seemed indicative of order. The absolute mean-residue
molar ellipticity at both 202 and 223 nm decreases with
growing chain length suggesting that the secondary structure
of longer peptide chains from 2 is destabilized. Similarly,
the CD spectra of oligomers (n ) 3, 6) of all (S)-3 with
terminal N-Boc and OEt groups in methanol also gave some
† Temple University.
‡ Merck Research Laboratories.
§ University of Pennsylvania.
| Department of Chemistry, University of Wisconsin-Madison.
⊥ Departments of Chemistry and Biochemistry, University of Wisconsin-
Madison.
(1) (a) Kakinoki, S.; Hirano, Y.; Oka, M. Polym. Bull. 2005, 53, 109–
115. (b) Cowan, P. M.; McGavin, S. Nature 1955, 176, 501–503. (c) Traub,
W.; Shmueli, U. Nature 1963, 198, 1165–1166.
(2) Abele, S.; Vogtli, K.; Seebach, D. HelV. Chim. Acta 1999, 82, 1539–
1558. It was known that peptoids form ordered secondary structures; see
refs 18-21 therein.
10.1021/ol1022917  2010 American Chemical Society
Published on Web 11/02/2010

indications of secondary structure. The mean residue molar
ellipticity of the hexamer at 230 nm is nearly three times
larger than the trimer indicating that in this case the
oxygens are not suitably oriented for nfπ* interactions with
the ꢀ-carboxyl side chains.5 A possible approach to solve
this problem is to limit the flexibility of the five-membered
ring, as in the methano-fused6 or ethano-bridged7 structures
6 or 7, or to control amide conformation by R,R-disubsti-
tution as in 8.8 Oligomers of 7 and 8 show evidence of order
secondary structure may be stabilized by the longer ꢀ-peptide
chain. Gellman4 independently investigated oligomers (n )
2-6) of all (S)-3 with terminal N-Boc and OBn groups in
methanol and observed a maximum ellipticity at 228 nm
when n ) 3-6. However, little evidence beyond CD spectra
was provided to show that these ꢀ-amino acid oligomers
adopted well-defined structures in solution.
in their CD spectra; a possible structure for oligomers of 7
has been calculated.
An alternative conformational restriction is the introduction
of a methano-bridge into an N-acyl-pyrrolidine ring to
maintain an idealized Cꢀ ring pucker, as in structure 9.9 The
bridge constrains two internal angles of the pyrrolidine (Φ
∼ -127° and θ ∼ 54°). Of the two remaining rotatable
bonds, the amide bond might be biased with the carbonyl
oriented either syn or anti to the bridgehead C1, so that ω ∼
180° or 0°. Thus, the major remaining rotational freedom
responsible for secondary structure would be from the
external C5-CO bond (Ψ). We believed that introducing
such a conformational constraint in ꢀ-proline oligomers could
provide entry into a well-defined and potentially useful
foldamer secondary structure.
A model was proposed for the oligomeric peptides of 2.2
It assumed an all-trans amide structure with a Φ angle of
-72° enforced by the pyrrolidine ring. An antiperiplanar
conformation around the C(R)-C(ꢀ) bond (θ ) 180°) was
chosen on the basis of this angle in a trifluoroacetate salt of
a trimer of 2 that contains amine and benzyl ester end groups.
However, it should be noted that the peptide salt crystallized
with four CF3COOH molecules and the carbonyls of the
amide bonds of the salt were hydrogen bonded to acid
molecules. A large angle, Ψ ) 180°, was chosen so that the
large substituents at C(R) were antiperiplanar to the large
substituents at the carbonyl C-atom. The outcome of this
somewhat speculative model is a right-handed 103 helix with
three pitches to bring residue (i + 10) above residue i. The
model suggests consecutive fully extended chain segments
[N-C(ꢀ)-C(R)-CO-N], which are twisted by -72°. The
structure of peptide oligomers of 3 was not modeled.
(R)-3-Carboxypyrrolidine 4 (PCA) might also be viewed
as a ꢀ-amino acid proline analogue in which the acid is
moved one atom away from the nitrogen. Gellman3 has used
the (S)-PCA enantiomer to search for order in pyrrolidine-
based oligomers (n ) 2-6). Normalized CD spectra in
methanol for oligomers 5 terminated by N-Boc and OBn
groups are most ordered when at the tetramer to hexamer
level with a minimum ellipticity at 214 nm. A possible
conformation has been modeled by calculation.4
In preparation for homooligomer formation, racemic acid
9 was resolved as its salt using (S)-(-)-R-methylbenzylamine
with THF as recrystallization solvent. The absolute config-
uration of the resolved (-) acid 9 was established as
(1S,4R,5R) by X-ray structure determination of the condensed
amide 10.
One of the difficulties in evaluating the order in homoo-
ligomers of pyrrolidine-based ꢀ-amino acids such as 2 or
PCA 4 is the number of degrees of freedom in the molecules.
There are no intramolecular hydrogen bonds to control
geometry, and unlike N-acyl prolines, the amide carbonyl
(5) (a) Shoulders, M. D.; Raines, R. T. Annu. ReV. Biochem. 2009, 78,
929–958. (b) Choudhary, A.; Gandla, D.; Krow, G. R.; Raines, R. T. J. Am.
Chem. Soc. 2009, 131, 7244–7246. See refs 3-6 therein.
(6) Medda, A. K.; Lee, H.-S. Synlett 2009, 921–924.
(7) Otani, Y.; Futaki, S.; Kiwada, T.; Sugiura, Y.; Muranaka, A.;
Kobayashi, N.; Uchiyama, M.; Yamaguchi, K.; Ohwada, Tetrahedron 2006,
62, 11635–11644.
(8) (a) Huck, B. R.; Fisk, J. D.; Guzei, I. D.; Carlson, H. A.; Gellman,
S. H. J. Am. Chem. Soc. 2003, 125, 9035–9037. (b) Huck, B. R.; Gellman,
S. H. J. Org. Chem. 2005, 70, 3353–3362.
(3) Huck, B. R.; Langenhan, J. M.; Gellman, S. H. Org. Lett. 1999, 1,
1717–1720.
(4) Sandvoss, L. M.; Carlson, H. A. J. Am. Chem. Soc. 2003, 125,
(9) Krow, G. R.; Lin, G.; Herzon, S. B.; Thomas, A. M.; Moore, K. P.;
Huang, Q.; Carroll, P. J. J. Org. Chem. 2003, 68, 7562–7564.
15855–15862.
Org. Lett., Vol. 12, No. 23, 2010
5439

To test for a conformational trans/cis bias of the amide
angle (ω), MPCA 9 was converted with Me3SiCHN2 into
its methyl ester 11 and then by protecting group exchange
into the isobutyryl amide 12. NMR analysis of monomer 12
indicated two H1 resonances in a 1:1 ratio indicating no
inherent bias for trans-/cis-amides. The dimer 13a was
prepared by cleavage of the N-Boc group from ester 11 with
HCl/dioxane and direct coupling of the resulting amine with
(-)-acid 9 using EDC and HOBt. Multiple conformations
for dimer 13a are possible. For each of two amide
N(sp2)-C(sp2) and two C5(sp3)-C(sp2) bonds there are,
respectively, two and three different staggered conformations.
In theory, there are at least 36 potential local minima. Despite
11 and the oligomers (n ) 2, 4, 6, 8) are shown in Figure
1A. The data for ꢀ-peptide concentration and number of
1
this, the H NMR spectrum of dimer 13a showed just two
major downfield H1 resonances. To facilitate the spectral
analysis of these major resonances, the terminal N-Boc group
of 13a was replaced with an N-isobutyryl substituent to give
13b. Expansion of the downfield H1 region of dimer 13b
indicated ten H1 resonances that individually integrate for
at least 3% of the total and that together account for 95% of
the H1 peaks; however, two major H1 resonances together
represent 62% of the H1 peaks. The key NMR resonances
associated with the major conformation of dimer 13b are
shown in Table 1. NOE data show that the isopropyl CHMe2
Figure 1. CD spectra for (A) 100 µM N-BOC-(MPCA)n-OMe in
methanol (25 °C). (B) 100 µM octamer 16a in water, nPrOH, and
mixtures of water/nPrOH (25 °C). Data are normalized for ꢀ-peptide
concentration and number of amides.
Table 1. Key Major NMR Resonances for Dimer 13b
amino acid units were normalized to facilitate comparisons
among oligomers of different lengths. The CD spectra of
tetramer, hexamer, and octamer have a strong negative molar
ellipticity at 198 nm and a weak positive at 212-215 nm.
The overlapping of spectra from monomer to octamer clearly
shows that substantial order begins to develop at the tetramer
level and is enhanced in longer oligomers. The positive molar
elipticity increases with increasing oligomer length and shifts
to slightly shorter wavelength.
CD spectra of the octamer 16a in water, nPrOH, and mixed
solvents of water/nPrOH in different ratios were collected
to study the solvent effects on conformation of the oligomer
(Figure 1B). From these spectra, it is clear that water has
little effect on the secondary structure of octamer. Because
the backbone of these peptides is tertiary amides, there are
no internal hydrogen bonds for water to disrupt.
The N-Boc-tetramer 14a formed a crystalline solid suitable
for X-ray analysis (Figure 2). Although a water molecule
attached to the C5-carbonyl oxygen (O2) of unit 2 alters the
Ψ angles predicted by calculations (lower picture in Figure
2), the tetramer does have a T4T4T4T4 arrangement that
places each carbonyl, including the terminal ester group, so
that alternate dipoles point in opposite directions (see dimer
13b above).
In solution, the N-Boc tetramer 14a exhibits four major
H1 peaks that represent 58-72% of a major conformation.
The NMR analysis of the N-isobutyryl tetramer 14b (Table
2) shows that the major (72%) structure can be described as
a T4T4T4T4 conformation with the final ester conformation
not determined by NMR (see Supporting Information).
proton
unit 1
2.52a
unit 2
carbon
unit 1
unit 2
CH
H1
H3
H3′
H4
-
31.9
60.1
47.7
5.10b c
4.88d
3.70
3.75
3.04
C1
C3
60.9
46.5
,
3.37
3.81
3.18
C4
40.4
41.3
a NOE cross-peak with H3,3′ of unit 1. b Coupled to C3 of unit 1 (HMBC).
c NOE cross-peak with H3,3′ of unit 2. d Coupled to C3 of unit 2.
is near the methylene H3,3′ protons of unit 1. The C3 carbon
associated with these H3,3′ protons (by HSQC) is coupled to
H1 of this first unit (by HMBC). Most importantly, the H1
proton of the first unit has an NOE cross-peak with the H3,3′
protons of the second unit, H(1,i)-H(3,i + 1). The H4
resonances for units 1 and 2 could be identified by their
W-plan coupling with H1 protons for each unit. We designate
this major dimer conformer as the T4T4 structure. The first
amide bond is trans; the carbonyl of unit 1 is directed toward
H4; and the second carbonyl is trans. To minimize dipole
repulsions, the ester carbonyl is assumed to be oriented
toward H4 (vide infra).
With evidence of some solution-phase order for the dimers
13a-13b in hand, the oligomers 14a-16a were prepared
(see Supporting Information). The CD spectra of monomer
5440
Org. Lett., Vol. 12, No. 23, 2010

COOMe configuration. A dependence of conformational
stabilization (n ) 2-8) on chain length also was noted for
oligomers of 7. Like the present (T4)n structures for
oligomers of 9, calculations for the tetramer of 7 predict Ψ
) 65° to be slightly favored.7 Calculations for oligomers 5
predict (C1)n conformations to be favored.4 Notably, the
(C1)4 and (T4)4 oligomers under discussion do have alternat-
ing anti orientations for their carbonyl dipoles in common.
There is indication of a major conformation in each of
our oligomers, as shown by the H1 region of each 1H NMR
spectrum (see Supporting Information). The crystal structure
of tetramer 14a, in silico methods, and the solution NMR
analyses of oligomers 13b-16b suggest [T4]n structures for
all amide bonds as the major contributors. The ability to have
a well-defined folding pattern in the absence of hydrogen
bonds or substantial electrostatic and hydrophobic interac-
tions should allow for the creation of foldamers with
interesting functions.11 Thus, the MPCA 9 may serve as a
useful addition to the arsenal of ꢀ-amino acid building
blocks.12 We are currently preparing C1- and C6-substituted
MetPyr acid oligomers that might adopt other interesting
conformations amendable to NMR analysis. For example,
although N-acyl-1-methyl-2-azabicyclo[2.1.1]hexanes are
restricted to the conformer with the carbonyl directed to the
C1 position,13 in oligomers of such substrates steric interfer-
ence is not likely to favor the Me(1,i)-H(3,i + 1) interactions
present in (T4)n conformations.
Figure 2. (Top) ORTEP drawing of tetramer 14a. (Bottom) Spartan
MMFF minimized (T4)n conformer of tetramer 14a.
Molecular mechanics geometry optimizations of 14b10 and
an N-acetyl tetramer 14c (see Supporting Information)
Table 2. Key Major NMR Resonances for Tetramer 14b
Acknowledgment. Financial support was provided by
NSF (CHE 0515635 and DGE-0841377) and the NIH
(AR044276). CD spectra were collected at the Biophysics
Instrumentation Facility established by Grants BIR-0-
9512577 (NSF) and S10 RR13790 (NIH). M.S. was sup-
ported by a Der Min Fan Fellowship, and M.D.S. was
supported by a USA Department of Homeland Security
Graduate Fellowship. We thank Rocky Chiang, Kevin
Cannon, Ram Edupuganti (TU), and Samuel H. Gellman
(UW-Madison) for helpful discussions.
proton
unit 1
unit 2
unit 3
unit 4
CH
H1
H3
2.52a
5.08b
3.35
-
4.95b
3.68
3.71
4.97b
3.57
3.74
4.87
3.58
3.70
H3′
3.77
a NOE with to H3,3′ of unit 1. b NOE to H3,3′ of the next unit.
Supporting Information Available: General experimental
and procedures for preparation of all new compounds, X-ray
diffraction analyses of amide 10 and tetramer 14a, spectral
assignments for tetramer 14b, a table of H1 resonances for
all oligomers, a calculated (T4)4 structure 14c, and copies
of reported 1H NMR and 13C NMR for all new compounds.
This material is available free of charge via the Internet at
http://pubs.acs.org.
indicate a preference for [T4]n conformations. The calculated
angles for 14b as a [T4]n conformation are ω ∼ 176-179°
and Ψ ∼ 58-64.5°. Importantly, in the favored calculated
structure H1 of the nth residue was in close proximity to
H3,3′ of the (n + 1) residue. Orientations in which the C5
carbonyl of unit i is oriented toward H1 of unit i + 1, or the
amide-carbonyl of any unit is oriented cis toward H3,3′ rather
1
OL1022917
than H1, are inconsistent with H NMR NOE correlations
linking H1 (unit i) with H3,3′ (unit i + 1).
(11) (a) Gellman, S. H. Acc. Chem. Res. 1998, 31, 173–180. (b) Hill,
D. J.; Mio, M. J.; Prince, R. B.; Hughes, T. S.; Moore, J. S. Chem. ReV.
2001, 101, 3893–4011. (c) Cheng, R. P.; Gellman, S. H.; DeGrado, W. F.
Chem. ReV. 2001, 101, 3219–3232. (d) Seebach, D.; Gardiner, J. Acc. Chem.
Res. 2008, 41, 1366–1375.
The CD spectra for the MPCA oligomers in Figure 1 are
similar to the characteristic concentration-independent CD
molar elipticities at 217 nm (maximum) and 198 nm
(minimum) found for oligomers of azabicycle 7 of the 2S-
(12) (a) Horne, W. S.; Gellman, S. H. Acc. Chem. Res. 2008, 41, 1399–
1408. (b) Seebach, D.; Beck, A. K.; Bierbaum, D. J. Chem. BiodiVersity
2004, 1, 1111–1239.
(10) (a) PC Spartan’04, Essential V2.0.0 Wavefunction, Inc.: Irvine,
CA 92612, 2004. (b) A rise/residue of 3.5Å, 2.4 residues/turn, and a pitch
of 8.4 Å can be approximated from the calculated structure.
(13) Malpass, J. R.; Patel, A. B.; Davies, J. W.; Fulford, S. J. Org. Chem.
2003, 68, 9348–9355.
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