A nonpeptidic reverse turn that promotes parallel sheet structure stabilized by C-H...O hydrogen bonds in a cyclopropane γ-peptide

Article abstract of DOI:10.1002/anie.200802648

(Chemical Equation Presented) A twist of fate: Parallel-turn linkers comprising an amino acid derived alcohol conjoined with an aromatic amide through a flexible linkage adopt reverseturn conformations. Cyclopropane tetra-and hexapeptide analogues form a C-H...O hydrogen-bond-stabilized parallel-sheet conformation (see scheme). NMR studies confirm the presence of hydrogen bonds in these structures.

Full text of DOI:10.1002/anie.200802648

Angewandte
Chemie
DOI: 10.1002/anie.200802648
Foldamers
A Nonpeptidic Reverse Turn that Promotes Parallel Sheet Structure
À
Stabilized by C H···O Hydrogen Bonds in a Cyclopropane g-Peptide**
Christopher R. Jones, M. Khurram N. Qureshi, Fiona R. Truscott, Shang-Te Danny Hsu,
Angus J. Morrison, andMartin D. Smith*
It is remarkable that compact folded proteins are constructed
from a small number of secondary structural elements such as
helices and sheets. Examination of model systems for these
structural modules has permitted understanding of the
features that regulate their assembly, independent of a
tertiary structural context. An extension of these principles
has led to the development of non-natural folded oligomers
(“foldamers”)[1] that populate distinct secondary and tertiary
structures (Figure 1).[2]
Amongst these folding backbones, parallel-[3] and anti-
[4]
parallel-sheet secondary structure has been demonstrated
in both b- and g-peptides,[5] and turn elements as simple as l-
dOrn (l-dornithine)have been demonstrated to nucleate
hairpin formation in organic and aqueous solutions.[6]
A
range of parallel-sheet models[7] have been delineated, as
exemplified by the d-Pro-DADME (d-proline-1,1-dimethyl-
1,2-diaminoethane)reverse turn, an elegant evolution of the
naturally inspired d-Pro-Xxx dipeptide (Xxx = any other
amino acid).[8] This material has been demonstrated to be
effective and versatile in promoting parallel-sheet structure in
a wide range of amide-linked strand sequences.[9] Here we
demonstrate that a reverse turn comprising an amino acid
derived alcohol conjoined with an aromatic amine can
promote parallel-sheet structure in a g-peptide sequence
À
designed to fold with the aid of C H···O hydrogen bonds.
Sheet construction is challenging as it requires the inter-
play of functional groups that are geometrically far apart in
primary structure. Hence, the employment of an appropriate
linking segment is of paramount importance. In designing this
parallel-turn motif (Figure 1), we reasoned that the incorpo-
ration of an ortho-amino phenol derivative would restrict the
y(i+2) torsion to angles consistent with natural b turns.[10] An
N-aryl amide proton would also possess a greater hydrogen-
bond-donor ability than a conventional amide, and the
presence of the aryl trifluoromethyl group may offer further
conformational control through acidification of the ortho
proton, facilitating its participation in a hydrogen bond with
the adjacent carbonyl group.[11] This additional hydrogen
bond has the potential to enhance the donor ability of this
amide further, through a cooperative mechanism.[12] We
considered that replacement of the amide linkage in the
rigid backbone of the Pro-Xxx model with an aryl ether could
offer sufficient conformational flexibility to enable favorable
hydrogen-bonding geometries to be populated; intramolecu-
lar hydrogen bonding offers a moderate driving force for
folding in an aprotic solvent. This motif also permits a range
of side chains derived from a-amino acids to be incorporated,
generating a simple and tunable alternative reverse-turn
linker.
Figure 1. Design of new parallel-turn linkers.
[*] C. R. Jones, M. K. N. Qureshi, F. R. Truscott, Dr. S.-T. D. Hsu,
Dr. M. D. Smith
Department of Chemistry, University of Cambridge
Lensfield Road, Cambridge CB2 1EW (UK)
Fax: (+44)1223-336-362
E-mail: mds44@cam.ac.uk
Dr. A. J. Morrison
Organon Research
Newhouse, Lanarkshire ML1 5SH (Scotland)
In a preliminary investigation to evaluate the turn-
forming propensities of this construct, we generated units
derived from l-valine (Val), l-tert-leucine, and l-proline
(Scheme 1). These units were assembled through an SNAr
reaction between the respective amino acid derived potas-
sium alkoxide (from 2 and 3)and commercially available
fluoroarene 1. Unblocking of the orthogonal N termini
(through transfer hydrogenation of the aryl nitro group and
subsequent removal of the alkyl N-Boc group, Boc = tert-
[**] We acknowledge funding from the Royal Society (M.D.S.), Organon/
EPSRC(C.R.J.), AstraZeneca (F.R.T.), the Pakistani HEC(M.K.N.Q.),
EPSRCNational Mass Spectrometry Centre, and an International
Human Frontier Science Program Long Term Fellowship LT00798/
2005 (S.T.D.H.). We would like to thank Dr. J. Davies for help with
X-ray crystallography.
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/anie.200802648.
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With the potential of these scaffolds to populate turn
conformations established, we decided to explore their utility
in a demanding scenario through the generation of a g-
peptide sheet using the valine-derived turn construct 4. We
have previously shown that cyclopropane g-peptides adopt
extended conformations in the solid state[15] and envisaged
that these materials could adopt sheet structures when
attached to an appropriate turn linker (Figure 3).[16]
Scheme 1. a) KHMDS, THF, 08C ; b) NH·HCOO, Pd/C, MeOH, 508C;
4
c) Ac2O, pyridine, DMAP; d) TFA/CH2Cl2, 08C ; e) AcO, pyridine,
2
DMAP. KHDMS=potassium hexamethyldisilazide, DMAP=N,N-
dimethyl-4-aminopyridine, TFA=trifluoroacetic acid.
butyloxycarbonyl), and attachment of minimal side chains (in
this case, acetyl groups)were accomplished without incident
to afford 6–8. The role of these model systems is to investigate
the turn-forming ability of this motif in the absence of
complicating effects from any side chains, and the conforma-
tion of these materials was investigated in both solution and
the solid state (Figure 2).[13]
Figure 3. Design rationale: from inter- to intramolecular hydrogen
bonds.
À
We reasoned that exploiting the intermolecular C H···O
hydrogen bonds present in oligomeric cyclopropane g-amino
[15]
acids 10 in the solid state
as stabilizing interactions in the
intramolecular manifold (as exemplified by 11)would be an
interesting test of our turn construct and of the potential role
[17]
À
of C H···O interactions
in unnatural sheet formation.
High-resolution protein structures have been shown to
contain sheet regions with short C H···O distances, and
NMR spectroscopy has provided direct evidence of hydrogen
[18]
À
Figure 2. X-ray structures of model turn constructs.
À
=
bonding across C H···O C in proteins by measuring through-
h3
bond
J
scalar couplings.[19] However, there are few
CaC’
examples of the exploitation of this interaction in the
development of novel folding backbones.[20] The absolute
configuration of proline in Pro-Xxx reverse turns can have a
profound effect on the propensity for sheet formation, and
hence we accessed both the d- and l-valine series of our
construct in this regard.[21] We were encouraged by the X-ray
structure[22] of an intermediate (in the d-Val series) 12 that
exhibited a turn structure stabilized by an intramolecular
bifurcated hydrogen bond that includes the CaH atom of the
cyclopropane group (Figure 4).
X-ray crystallography of 6 and 7 (recrystallized from
CH2Cl2)revealed a conformation characterized by both inter-
and intramolecular hydrogen bonding. We were unable to
obtain suitable crystals of l-Pro N-acetate compound 8 but
were able to solve the structure of N-Boc derivative 9. For 6,
the intramolecular donor–acceptor distances are favorable
a
=
À
=
=
(C O···H N: 2.2 Š and C O···C H: 2.5 Š)though the C
À
O···H N angle deviates significantly from planarity (1008).
The l-tert-leucine-derived unit 7 has a similar geometry in the
solid state, and the l-Pro derivative 9 adopts a comparable
hydrogen-bonded turn geometry. The properties of these
materials were also investigated by FTIR and 1H NMR
spectroscopy in nonpolar solvents. Titration experiments
(1 mm in CDCl3, 298 K)indicate in both cases that the
dependency of the chemical shift of the aryl amide proton on
the concentration of DMSO is ten times lower than that of the
alkyl amide proton, which is consistent with its involvement in
intramolecular hydrogen-bonded conformations. The FTIR
spectrum of 6 (1 mm in CH2Cl2, 298 K)shows two peaks in the
The cyclopropane CaH···O separation (2.3 Š)is consistent
with similar interactions[23] observed in natural[24] and non-
natural sequences.[25] The preference for linearity across the
CaH···O interaction clearly influences the conformation
adopted by the cyclopropyl side chain. In this hydrogen-
À1
À1
À
N H stretch region at 3434 cm (sharp)and 3333 cm
À
(broad), which we ascribe to non-hydrogen-bonded N H
[13]
À
and internally hydrogen-bonded N H groups respectively.
Similar data were recorded for the l-tert-leucine-derived
material 7 (N H stretch region: 3439 cm and 3334 cmÀ1),
À1
À
consistent with an analogous hydrogen-bonded conforma-
À
Figure 4. X-ray structure of 12 showing a C H···O stabilized turn
conformation.
tion.[14]
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Table 1: Chemical shifts of amide and CaH cyclopropane protons.[a]
bonded conformation, the side chains attached to the reverse
turn in 12 are oriented in different directions, which has
significant implications for the ability of materials based on
this specific diastereoisomeric motif to populate extended
parallel-sheet conformations. Close contacts such as these
may be regarded as a consequence or a determinant of the
observed geometry in molecular crystals, and hence it is
important to gain further evidence of their role in determining
conformation.[26]
NHA
NHB
NHC
NHD
C-2C
C-2D
C-2E
C-2F
13
14
15
16
8.71
8.71
8.70
8.75
5.83
5.68
5.96
5.81
–
–
6.54
6.54
–
–
6.11
6.97
2.03
1.98
1.89
1.98
1.77
1.38
1.58
1.50
–
–
1.37
1.68
–
–
1.37
1.60
[a] Conditions: 700 MHz, 3 mm in CDCl3 (referenced to 7.27 ppm),
298 K.
We envisaged that CaH···O interactions such as these
could stabilize a parallel sheet in solution, and hence
tetrapeptide analogues 13 and 14, and hexapeptide analogues
15 and 16 were synthesized, and their conformational
preferences examined by NMR spectroscopy in organic
solvents (Figure 5).
In order to probe the individual hydrogen-bonding
interactions in materials 13–16, we carried out a series of
variable-temperature and solvent-dependent experiments.[14]
Solvent titration studies demonstrate that only the cyclo-
propane CaH and amide protons possess a chemical shift
dependence on DMSO concentration, which is consistent
with their involvement in hydrogen bonding. For tetrapeptide
analogue 14, NHA has an approximate sevenfold lower
chemical shift dependency than NHB with respect to DMSO
concentration, and a similar trend is seen for the CaH at C-2C
when compared to that at C-2D. For 16, these experiments
demonstrate that NHA and NHD are solvent-protected
relative to NHB and NHC whilst for 15, the chemical shift
dependency on DMSO concentration of NHB, NHC, and NHD
is significantly higher than that of NHA. This is consistent with
the NOE data for 15 that indicate close contacts at the hairpin
end of the molecule only.
Temperature coefficients were also determined for 13–
16.[13,14,28] Compounds 13 and 14, which have similar con-
formational profiles, possess similar temperature coefficient
data. In contrast, 15 and 16, which differ in conformation at
the N termini, display significantly different temperature-
related data that we believe reflect unfolding and conforma-
tional averaging.[29]
Figure 5. Selected interresidue NOE correlations for valine-derived
sheets (700 MHz, 3 mm in CDCl3 (referenced to 7.27 ppm), 298 K).
When considered together, this evidence suggests that
tetrapeptide analogues 13 and 14 populate well-defined turn
conformers stabilized by intramolecular hydrogen bonds in
which both aromatic amide and the cyclopropane CaH
protons are implicated as hydrogen-bond donors. In contrast,
15 and 16 (which differ only in the absolute configuration of
the asymmetric center on the reverse-turn)populate signifi-
cantly different conformations. For 15, the turn structure is
conserved at the hairpin end of the molecule (consistent with
the X-ray structure of 12)but there is little evidence of long-
range order, whilst 16 populates an extended sheetlike
conformation stabilized by a series of hydrogen bonds.
In summary, we have generated a new nonpeptidic reverse
turn that populates a hairpin conformation in the solid state
and in solution. Exploitation of the well-defined conforma-
tional preferences of this construct enabled the generation of
sheetlike materials that populate hydrogen-bond-stabilized
conformations. Both solid-state and solution data indicate
For these materials, dilution experiments confirmed that
no intermolecular aggregation was occurring at concentra-
tions below 3 mm, and a series of 2D experiments permitted
assignment of all spin systems. Stereospecific assignment of
the cyclopropyl methylene protons was achieved unambigu-
ously for almost all spin systems; this is important as specific
nuclear Overhauser effect (NOE)signals from these protons
are key in defining turnlike conformers in solution. For the
tetrapeptide analogues 13 and 14, cross-strand NOE correla-
tions are observed between nonadjacent cyclopropane rings,
consistent with population of
a hairpin conformation
(Figure 5). The l-Val-derived hexapeptide 16 possesses key
NOE correlations involving the terminal cyclopropane resi-
dues, indicative of the population of a sheet conformer whilst
in the d-Val-derived analogue 15 these key NOE correlations
1
are absent. H NMR chemical shift data for 16 (Table 1)is
consistent with the involvement of NHD and NHA in hydrogen
bonds (both are deshielded), whilst for 15 this effect is
exhibited only by NHA. For 16, the cyclopropane CaH signal
at C-2C is deshielded relative to that at C-2D, as expected in its
role as a hydrogen-bond donor.[27] Similar shift patterns are
seen for 13–15.
significant C H···O hydrogen bonding in these materials,
À
which is consistent with the postulated structural role of this
interaction in proteins. We are currently exploring the utility
À
of the C H···O hydrogen bond as a tool for conformational
control in small molecules in aqueous and organic solvents.
Our work thus far augurs well for future application of these
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[16] For
a cyclopropyl b-amino acid that populates a helical
scaffolds in the generation of well-defined materials for
conformation see: S. De Pol, C. Zorn, C. D. Klein, O. Zerbe,
O. Reiser, Angew. Chem. 2004, 116, 517; Angew. Chem. Int. Ed.
2004, 43, 511.
catalysis and recognition.
Received: June 5, 2008
Published online: July 30, 2008
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