A new artificial β-sheet that dimerizes through parallel β-sheet interactions

Article abstract of DOI:10.1021/ol802993v

This paper introduces a chemical model of aβ-sheet that dimerizes through parallelβ-sheet interactions in CDCI₃ solution. The model consists of two C-terminally linked dipeptides connected to a molecular template. ¹H NMR studies esta

Full text of DOI:10.1021/ol802993v

ORGANIC
LETTERS
2009
Vol. 11, No. 4
1003-1006
A New Artificial ꢀ-Sheet That Dimerizes
through Parallel ꢀ-Sheet Interactions
Sergiy Levin and James S. Nowick*
Department of Chemistry, UniVersity of California, IrVine,
IrVine, California 92697-2025
jsnowick@uci.edu
Received December 31, 2008
ABSTRACT
This paper introduces a chemical model of a ꢀ-sheet that dimerizes through parallel ꢀ-sheet interactions in CDCl3 solution. The model consists
of two C-terminally linked dipeptides connected to a molecular template. 1H NMR studies establish the ꢀ-sheet folding and dimerization of the
model system. This system corroborates that linking two peptide strands and blocking one edge of the assembly creates soluble, easy-to-
study systems that participate in the types of interactions that occur widely in peptide and protein aggregates.
Intermolecular parallel ꢀ-sheet interactions occur widely in
peptide and protein aggregation and are important in Alzhe-
imer’s and a variety of other neurodegenerative disorders.1
These noncovalent interactions are difficult to study because
they often occur in an uncontrolled fashion and lead to
insoluble aggregates. We recently reported the first well-
defined chemical model system with which to study inter-
molecular parallel ꢀ-sheet formation.2 Here, we report a
second chemical model system that forms well-defined
dimers through parallel ꢀ-sheet formation.3
In our first system, we achieved dimer formation by
N-terminally linking two short peptides with a succinic acid
unit and blocking one of the exposed hydrogen-bonding
edges with a molecular template. In the new system, we
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Shewmaker, F.; Wickner, R. B.; Tycko, R Proc. Natl. Acad. Sci. U.S.A.
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10.1021/ol802993v CCC: $40.75
Published on Web 01/27/2009
2009 American Chemical Society

Scheme 1. Building Blocks of Artificial ꢀ-Sheet 2
Artificial ꢀ-sheet 2 consists of two C-terminally linked
dipeptide strands that are blocked along one edge by a
hydrogen-bonding template. The dipeptide strands are linked
with ethylenediamine, and the template comprises 5-ami-
noanisic acid oxalamide and valine. The template is con-
nected to one of the dipeptide strands by a turn unit
composed of δ-linked ornithine.4 Scheme 1 illustrates these
components. The exposed edge of the linked dipeptide
strands of 2 presents six alternating hydrogen-bond donor
and acceptor groups that participate in the formation of the
hydrogen-bonded dimer. We incorporated the nonpolar
amino acids leucine and valine in the peptide strands and
template to allow studies in chloroform, a solvent that
supports hydrogen bonding.
Artificial ꢀ-sheet 2 was assembled from 5-aminoanisic
oxalamide template 3 and ethylenediamine-linked peptide 4
Scheme 2. Assembly of Artificial ꢀ-Sheet 2
Figure 1. Artificial ꢀ-sheets 1 and 2 and corresponding parallel
ꢀ-sheet dimers 1·1 and 2·2.
achieve dimer formation by C-terminally linking two short
peptides with an ethylenediamine unit and blocking one of
the exposed hydrogen-bonding edges. Figure 1 illustrates the
structure of the two systems, artificial ꢀ-sheets 1 and 2,
respectively, and the corresponding hydrogen-bonded dimers
1·1 and 2·2.
1004
Org. Lett., Vol. 11, No. 4, 2009

Figure 2. Key NOEs observed in artificial ꢀ-sheet 2. A: Red arrows
represent intermolecular NOEs; blue arrows represent intramolecular
NOEs. B: Newman projection with intramolecular NOEs (red
arrows).
using standard solution-phase peptide synthesis techniques
(Scheme 2). Peptide-coupling reactions were performed with
HCTU and 2,4,6-collidine or DIPEA.5 The 5-aminoanisic
oxalamide template was prepared from Fmoc-protected
5-aminoanisic acid.6 Ethylenediamine-linked peptide 4 was
prepared
from
Boc-protected
ethylenediamine
(BocNHCH2CH2NH2) by standard solution-phase Boc- and
Cbz-based peptide synthesis procedures.
1H NMR studies establish the folding and dimerization of
artificial ꢀ-sheet 2 in CDCl3 solution. The proton resonances
of artificial ꢀ-sheet 2, although broadened at ambient
temperature, are sufficiently sharp at 268 K and 800 MHz
to be assigned definitively by COSY, TOCSY, and NOESY
techniques. Consistent with a hydrogen-bonded ꢀ-sheet
structure, the amide NH resonances are shifted downfield
Figure 3. Selected NOEs in artificial ꢀ-sheet 2. 800 MHz NOESY
spectrum recorded at 4 mM and 268 K in CDCl3 with a 400-ms
mixing time. Intramolecular NOEs are shown in blue; intermo-
lecular NOEs are shown in red.
(4) (a) Nowick, J. S.; Lam, K. S.; Khasanova, T. V.; Kemnitzer, W. E.;
Maitra, S.; Mee, H. T.; Liu, R. J. Am. Chem. Soc. 2002, 124, 4972–4973.
(b) Nowick, J. S.; Brower, J. O. J. Am. Chem. Soc. 2003, 125, 876–877.
(5) (a) Pon, R. T.; Yu, S.; Sanghvi, Y. S. J. Org. Chem. 2002, 67, 856–
864. (b) Marder, O.; Albericio, F. Chim. Oggi 2003, 21, 35–40.
(6) Green, J. J. Org. Chem. 1995, 60, 4287–4290.
compared to non-hydrogen-bonded amide resonances and the
amino acid R-proton resonances are shifted downfield
Org. Lett., Vol. 11, No. 4, 2009
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3D) and H6 of the template and Hen1 of the linker (Figure
3C). Noteworthy NOEs associated with parallel ꢀ-sheet
dimerization include NOEs between Leu1 R and Leu2 ꢀ and
between Leu3 R and Orn ꢀ (Figure 3A). Additional note-
worthy NOEs associated with parallel ꢀ-sheet dimerization
include NOEs between Leu2 NH and Leu1 R and between
Orn NH and Leu3 R (Figure 3B).
The present system is complementary to the original
system in that the peptide strands in 2 are C-terminally
linked, while those in 1 are N-terminally linked. Thus, this
system offers a new topology for dimer formation and
provides a distinct new class of dimers. This new system
also demonstrates the generality of the approach to achieving
dimerization through parallel ꢀ-sheet interactions by linking
two peptide strands and blocking one of the exposed edges
with a molecular template. Figure 4 provides cartoons that
show the structural relationship between the dimers of these
two systems.
Figure 4. Schematic representations of parallel ꢀ-sheet dimers 1·1
(A) and 2·2 (B).
compared to random coil values.7 The ornithine δ-protons
show significant magnetic anisotropy (1.06 ppm) and strong
NOE signals with the ornithine R-proton, reflecting a turn
conformation of the ornithine side chain.4
The 800 MHz NOESY studies definitively establish the
formation of a well-defined and folded hydrogen-bonded
dimer in CDCl3 solution. Artificial ꢀ-sheet 2 shows a rich
array of intramolecular NOEs characteristic of antiparallel
ꢀ-sheet folding and intermolecular NOEs characteristic of
parallel ꢀ-sheet dimerization.8 Figure 2 illustrates these
NOEs. Figure 2A shows the overall structure of the dimer,
while Figure 2B represents half of the 2-fold symmetrical
parallel ꢀ-sheet dimerization interface as a Newman projec-
tion. Intramolecular NOEs associated with folding are
represented by blue arrows, while intermolecular NOEs
associated with dimerization are represented by red arrows.
Figure 3 shows key regions of the NOESY spectrum.
Noteworthy NOEs associated with antiparallel ꢀ-sheet fold-
ing include strong NOEs between Leu2 R and Val R (Figure
In future studies, we will use systems such as these to
explore noncovalent interactions among parallel ꢀ-sheets.
These types of systems are ideally suited to such studies,
because they do not form uncontrolled aggregates, like simple
peptides. We further anticipate applying the design principles
established through these studies to create systems that
participate in parallel ꢀ-sheet interactions in aqueous solution
and using these systems to antagonize peptide and protein
aggregation.
Acknowledgment. We thank the NIH (GM-49076) for
grant support and Dr. Evgeny Fadeev (UCI) for collecting
800 MHz NMR data.
(7) (a) Wishart, D. S.; Sykes, B. D.; Richards, F. M. J. Mol. Biol. 1991,
222, 311–333. (b) Wishart, D. S.; Sykes, B. D.; Richards, F. M. Biochemistry
1992, 31, 1647–1651.
Supporting Information Available: Synthetic procedures
and NMR spectroscopic data. This material is available free
of charge via the Internet at http://pubs.acs.org.
(8) (a) Wuthrich, K. NMR of Proteins and Nucleic Acids; Wiley: New
York, 1986, 125129. (b) Nowick, J. S. Org. Biomol. Chem. 2006, 4, 3869–
3885.
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