Introduction of a triazole amino acid into a peptoid oligomer induces turn formation in aqueous solution

Article abstract of DOI:10.1021/ol070817y

Peptoids are a non-natural class of oligomers that are composed of repeating N-substituted glycine units and are capable of folding into helices that mimic peptide structure and function. In this letter, we report the concise synthesis of a 1,5-substitute

Full text of DOI:10.1021/ol070817y

ORGANIC
LETTERS
2007
Vol. 9, No. 12
2381-2383
Introduction of a Triazole Amino Acid
into a Peptoid Oligomer Induces Turn
Formation in Aqueous Solution
Jonathan K. Pokorski,§ Lisa M. Miller Jenkins,† Hanqiao Feng,‡
Stewart R. Durell,† Yawen Bai,‡ and Daniel H. Appella*,§
Laboratory of Bioorganic Chemistry, NIDDK, National Institutes of Health, DHHS,
Bethesda, Maryland 20892, Laboratory of Cell Biology, NCI, National Institutes
of Health, DHHS, Bethesda, Maryland 20892, and Laboratory of Biochemistry
and Molecular Biology, NCI, National Institutes of Health, DHHS,
Bethesdsa Maryland 20892.
appellad@niddk.nih.goV
Received April 5, 2007
ABSTRACT
Peptoids are a non-natural class of oligomers that are composed of repeating N-substituted glycine units and are capable of folding into
helices that mimic peptide structure and function. In this letter, we report the concise synthesis of a 1,5-substituted triazole amino acid (Tzl)
and its subsequent incorporation into a short peptoid. The Tzl amino acid was shown to induce turn formation in aqueous solution, thus
expanding the structural repertoire available to peptoid chemists.
Peptoids are a class of synthetic oligomers that consist of
repeating N-substituted glycine units and are capable of
folding into discrete structures.1 Peptoids bearing R-branched
side chains can fold into helices resembling the polyproline
type I (PPI) helix2 and are attractive pharmaceutical candi-
dates because of their enhanced proteolytic stability3 and
cellular uptake.4 In this context, peptoids’ propensity for helix
formation has been used to develop mimics of the antibacte-
rial magainin peptides5 and lung surfactant proteins.6 More
recently, macrocyclic peptoids were designed to mimic â-turn
formation in organic solvents.7 Expanding the structural
repertoire of peptoids to include turns, loops, and sheets
should aid the development of biologically active derivatives
where such motifs are relevant. Progress in the development
of noncyclic peptoid structure has shown that certain
nonamers can fold into loops in organic solvent,8 which can
be stabilized through the introduction of electron-poor
§ Laboratory of Bioorganic Chemistry.
† Laboratory of Cell Biology.
‡ Laboratory of Biochemistry and Molecular Biology.
(1) Simon, R. J.; Kania, R. S.; Zuckermann, R. N.; Huebner, V. D.;
Jewell, D. A.; Banville, S.; Ng, S.; Wang, L.; Rosenberg, S.; Marlowe, C.
K.; Spellmeyer, D. C.; Tan, R.; Frankel, A. D.; Santi, D. V.; Cohen, F. E.;
Bartlett, P. A. Proc. Natl. Acad. Sci. U.S.A. 1992, 89, 9367-9371.
(2) (a) Armand, P.; Kirshenbaum, K.; Goldsmith, R. A.; Farr-Jones, S.;
Barron, A. E.; Truong, K. T.; Dill, K. A.; Mierke, D. F.; Cohen, F. E.;
Zuckermann, R. N.; Bradley, E. K. Proc. Natl. Acad. Sci. U.S.A. 1998, 95,
4308-4314. (b) Wu, C. W.; Sanborn, T. J.; Huang, K.; Zuckermann, R.
N.; Barron, A. E. J. Am. Chem. Soc. 2001, 123, 6778-6784. (c) Wu, C.
W.; Sanborn, T. J.; Zuckermann, R. N.; Barron, A. E. J. Am. Chem. Soc.
2001, 123, 2958-2963.
(4) Kwon, Y.; Kodadek, T. J. Am. Chem. Soc. 2007, 129, 1508-1509.
(5) Patch, J. A.; Barron, A. E. J. Am. Chem. Soc. 2003, 125, 12092-
12093.
(6) Seurynck, S. L.; Patch, J. A.; Barron, A. E. Chem. Biol. 2005, 12,
77-88.
(7) (a) Nnanabu, E.; Burgess, K. Org. Lett. 2006, 8, 1259-1262. (b)
Shin, S. B. Y.; Yoo, B.; Todaro, L. J.; Kirshenbaum, K. J. Am. Chem. Soc.
2007, 129, 3218-3225.
(3) Miller, S. M.; Simon, R. J.; Ng, S.; Zuckermann, R. N.; Kerr, J. M.;
Moos, W. H. Drug DeV. Res. 1995, 35, 20-32.
10.1021/ol070817y This article not subject to U.S. Copyright. Published 2007 by the American Chemical Society
Published on Web 05/17/2007

aromatic residues.9 Aqueous conditions, alternatively, induce
a conformational change favoring the PPI helix owing to
the necessity of hydrogen bonding between the peptoid
termini for loop formation.
The key synthetic step in the sequence is the Huisgen
cycloaddition that yields bicyclic triazole 2 as a single
regioisomer. The lactam is then opened under strongly acidic
conditions followed by Fmoc protection of the primary amine
to yield Tzl monomer 3 in good yield. We anticipate that
Tzl will prove useful for a number of studies that require
the use of a ring-constrained amino acid.
The Tzl monomer was readily incorporated into peptoid
oligomers using a modified version of the submonomer
synthesis.15 Peptoids were designed so that the Tzl monomer
was located in an analogous position to the turn region of a
peptide in a hairpin conformation. The folded conformation
of Tzl peptoids was initially assessed using circular dichro-
ism. Peptoid pentamer 4 showed both a global minimum at
200 nm, and a local minimum at 220 nm (Figures 1 and 2).
Recent research has begun to elucidate the rules for hairpin
formation in peptides.10 Conformational constraint about the
loop region has proven to be a powerful method for enforcing
a hairpin structure in model peptides11 and has been extended
to the design of hairpin mimics of protegrins.12 However,
turn constraints have not yet been applied in peptoid design.
We hypothesized that the incorporation of a 1,5-substituted
triazole amino acid (3) into peptoid oligomers would induce
turn formation in a peptoid backbone. The triazole introduces
a constraint that is geometrically similar to a cis double bond,
while acting as an amide isostere with respect to polarity.
Furthermore, this amino acid would not disrupt the registry
of the peptoid, maintaining molecular spacing equivalent to
two peptoid monomers. In this communication we report the
synthesis of a 1,5-substituted triazole amino acid (Tzl) and
its effects upon the secondary structure of short peptoids.
The most common method for accessing the triazole
framework is the copper-catalyzed Huisgen cycloaddition.13
This methodology, though, was not suitable for our needs
because the favored product is the 1,4-substituted regioiso-
mer. There are few methods in the literature available to
access the 1,5-substitution pattern, but those proved either
unreliable or costly because of the use of organometallic
reagents.14 Thus, we developed a gram-scale synthesis that
utilizes a ring-constrained Huisgen cycloaddition to introduce
the 1,5-substitution pattern (Scheme 1). Synthesis of the Tzl
Figure 1. Peptoid side-chain structures and nomenclature.
Previous work with peptoids has attributed the negative peak
at 200 nm to a loop structure, while the minimum at 220
nm suggests a helical conformation.8 We speculated that 4
was in conformational equilibrium between two folded states.
This hypothesis was confirmed by the observation of two
equally intense sets of resonances in TOCSY spectra. To
Scheme 1. Tzl Synthesis
monomer is accomplished in three steps, with no chromato-
graphic purification required. Propargyl amine is first acy-
lated using bromoacetyl chloride. The resulting bromoacet-
amide (1) is converted to an azide upon treatment with NaN3.
(8) Huang, K.; Wu, C. W.; Sanborn, T. J.; Patch, J. A.; Kirshenbaum,
K.; Zuckermann, R. N.; Barron, A. E.; Radhakrishnan, I. J. Am. Chem.
Soc. 2006, 128, 1733-1738.
(9) Gorske, B. C.; Blackwell, H. E. J. Am. Chem. Soc. 2006, 128, 14378-
14387.
(10) For a discussion of recent model hairpin systems: Hughes, R. M.;
Waters, M. L. Curr. Opin. Struct. Biol. 2006, 16, 514-524.
(11) (a) Haque, T. S.; Gellman, S. H. J. Am. Chem. Soc. 1997, 119,
2303-2304. (b) Grison, C.; Coutrot, P.; Geneve, S.; Didierjean, C.; Marraud,
M. J. Org. Chem. 2005, 70, 10753-10764.
(12) Lai, J. R.; Huck, B. R.; Weisblum, B.; Gellman, S. H. Biochemistry
2002, 41, 12835-12842.
Figure 2. CD spectra of peptoid oligomers 4-7. Peptoid sequences
are as follows. 4: NsceNspeTzlNspeNae, 5: NsceNipNspeTzlNsnp-
NsbNae, 6: NsceNipNspeTzl1,4NsnpNsbNae, 7: NsceNipNspeSar-
SarNsnpNsbNae.
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Org. Lett., Vol. 9, No. 12, 2007

Figure 3. Structure of peptoid 5: (a) chemical structure of 5 showing long-range NOEs and side-chain nomenclature; (b) ensemble of 9
modeled structures of the major conformation of 5 derived from the NOE restraints. The role of the Tzl unit is seen in the upper right
portion of the figure. Average atomic rmsd’s: all atoms, 5.26 Å; backbone turn atoms, 2.55 Å. The mean violation per structure for 26
restraints is 8.3 Å. All NMR experiments were performed in aqueous buffer; 10 mM sodium phosphate, pH 7.0.
further bias the peptoid toward a hairpin conformation, two
hydrophobic residues were incorporated in peptoid 5 (Figures
2 and 3) to encourage hydrophobic collapse as seen in hairpin
peptides.16 The CD spectrum of this peptoid showed only a
single intense global minimum at 200 nm. Subsequent control
experiments where the Tzl unit was replaced with the
analogous 1,4-substituted triazole (Tzl1,4) (6) or with sar-
cosine (7) maintained a local minimum at ∼220 nm,
indicating that the Tzl unit induces a unique fold.
Peptoid 5 was further characterized by NMR spectroscopy.
The 2D TOCSY spectrum of 5 in aqueous solution produced
a high quality spectrum with well-dispersed resonances
corresponding to a single peptoid conformation. Assignments
were confirmed and distance constraints were obtained from
a 2D ROESY spectrum. Refinement yielded a major and
minor conformation that both converged to show a compact
hairpin structure, resembling a peptide hairpin (Figure 3).
The major structure is characterized by a highly conserved
turn region and flexible termini. The Tzl residue initiates a
turn in the backbone, of which the carbonyl carbons of Tzl
(C18) and Nsb (C9) represent the ends of a 10-atom turn.
These carbon atoms are ∼5 Å apart, well within the 7 Å
maximum prescribed to peptide turns,17 suggesting that the
backbone geometry is a competent turn mimic. In all
solutions, the N- and C-termini maintain a close proximity
that may be facilitated by a salt bridge or hydrogen bond.
The termini though, are inherently flexible and can orient
themselves either above or below the plane of the turn region.
The findings reported here represent the first account of
hairpin formation by a peptoid in aqueous solution. The
synthesis and subsequent incorporation of the Tzl monomer
in concert with principles derived from model hairpin
peptides were critical to successful peptoid design. These
results have major implications toward the de novo design
of functional nonnatural polymers and effectively extend the
range of secondary structures available to the peptoid
community.
Acknowledgment. This work was supported by the
National Institutes of Health, NIDDK intramural research
program. The authors would like to thank Professor Karl
Scheidt for helpful discussions and Dr. John Lloyd of the
proteomics and mass spectrometry facility at the NIH for
collection of mass spectra.
Supporting Information Available: Detailed procedures
for synthesis of all compounds, CD studies, and NMR
stucture determination. NMR TOCSY and ROESY data for
peptoid 5 are also present, along with information on the
minor peptoid structural conformation. This material is
available free of charge via the Internet at http://pubs.acs.org.
(13) Rostovstev, V. V.; Green, L. G.; Fokin, V. V.; Sharpless, K. B.
Angew. Chem., Int. Ed. 2002, 41, 2596-2599.
(14) (a) Krasinski, A.; Fokin, V. V.; Sharpless, K. B. Org. Lett. 2004, 6,
1237-1240. (b) Zhang, L.; Chen, X.; Xue, P.; Sun, H. H.; Williams, I. D.;
Sharpless, K. B.; Fokin, V. V.; Jia, G. J. Am. Chem. Soc. 2005, 127, 15998-
15999.
(15) Zuckermann, R. N.; Kerr, J. M.; Kent, S. B. H.; Moos, W. H. J.
Am. Chem. Soc. 1992, 114, 15998-15999.
(16) Maynard, A. J.; Sharman, G. J.; Searle, M. S. J. Am. Chem. Soc.
1998, 120, 1996-2007.
OL070817Y
(17) Ball, J. B.; Hughes, R. A.; Alewood, P. F.; Andrews, P. R.
Tetrahedron 1993, 49, 3467-3478.
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