An activated building block for the introduction of the histidine side chain in aliphatic oligourea foldamers

Article abstract of DOI:10.1016/j.tet.2011.11.066

A new N-Boc-protected monomer for the synthesis of oligourea foldamers containing the (1H-imidazolyl-4yl)methyl side chain of histidine, has been prepared in seven steps from Trt-His(τ-Trt)-OMe. This protecting group combination on histidine was found to

Full text of DOI:10.1016/j.tet.2011.11.066

Tetrahedron 68 (2012) 4492e4500
Contents lists available at SciVerse ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
An activated building block for the introduction of the histidine side chain
in aliphatic oligourea foldamers
,
y
a
,
,
Yella-Reddy Nelli a , Celine Douat-Casassus y, Paul Claudon a b, Brice Kauffmann c, Claude Didierjean d,
ꢀ
,
Gilles Guichard a
*
a
ꢀ
ꢀ
Universite de BordeauxdCNRS UMR5248, Institut Europeen de Chimie et Biologie, CBMN, 2 rue Robert Escarpit, Pessac 33607, France
b
c
ꢀ
ꢀ
CNRS, Institut de Biologie Moleculaire et Cellulaire, Laboratoire d’Immunologie et Chimie Therapeutiques, Strasbourg 67000, France
ꢀ
ꢀ
Universite de BordeauxdCNRS UMS3033, Institut Europeen de Chimie et Biologie, 2 rue Robert Escarpit, Pessac 33607, France
CRM2, UMR-CNRS 7036, Groupe Biocristallographie, Universite Henri Poincare, BP 239, Vandoeuvre 54506, France
d
ꢀ
ꢀ
a r t i c l e i n f o
a b s t r a c t
Article history:
A new N-Boc-protected monomer for the synthesis of oligourea foldamers containing the (1H-imida-
Received 28 September 2011
Received in revised form 3 November 2011
Accepted 23 November 2011
Available online 29 November 2011
zolyl-4yl)methyl side chain of histidine, has been prepared in seven steps from Trt-His(s-Trt)-OMe. This
protecting group combination on histidine was found to be critical to ensure efficient access to the
requisite activated building block. This new derivative, suitable for solid phase synthesis, expands the
current arsenal of building blocks with proteinogenic side chains useful for the design of peptidomimetic
oligourea foldamers.
Dedicated to Professor Dieter Seebach on
the occasion of his 75th birthday
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Histidine
Succinimidyl carbamate
Oligourea
Foldamer
Solid phase synthesis
Imidazole
1. Introduction
synthetic peptides, containing a
effective organocatalysts for various asymmetric transformations.8
Furthermore, His-rich -peptide sequences have also found in-
teresting biotechnological and biological applications. This includes
His-tags for protein purification,9 metal-dependent cell-penetrat-
ing peptides for drug-delivery purposes10 and cationic amphipathic
peptides for gene transfer.11
Synthetic foldamers (i.e., non-natural oligomers that fold into
well-defined structures), owing to their structural predictability
and large contact areas, represent privileged scaffolds to orient
recognition motifs in space and interact with a variety of targets
(metals, small molecules, biopolymers).12 Thus, sets of building
p-methylated imidazole moiety, as
Histidine, with its (1H-imidazolyl-4yl)methyl side chain is a re-
markable amino acid in many ways. Besides its intrinsic basicity,
crucial to the catalytic properties of many enzymes, the ability of
the imidazole ring to bind and coordinate metal ions is widely used
to stabilize tertiary folds and helical structures1 (e.g., zinc fingers
with Cys2His2 tetrahedral Zn(II) site2). Not surprisingly, His residues
a
have been incorporated in a variety of
structural motifs (single
a-peptide sequences and
a
-helices,3 -helical bundles,4 helix-loop-
a
helix,5 bba tertiary folds6) designed de novo to mimic protein metal
binding sites and/or active sites for catalysis. With the aim to re-
produce the microenvironment of enzyme catalytic pockets, Rey-
mond and co-workers have developed artificial catalytic
architectures based on His-rich peptide dendrimers.7 Using a bio-
mimetic approach, Miller and co-workers engineered small
blocks equivalent to
a-amino acids, i.e., bearing requisite in-
formation for folding and diversity in side chains are of particular
interest in the context of foldamer-based recognition. However,
examples of peptidomimetic foldamers with His-type side chains at
their surface are scarce. This can be ascribed in part to synthetic
difficulties created by the basic and nucleophilic 1H-imidazole ring.
The preparation of N-Fmoc-protected
mologues of histidine (in eight and ten steps, respectively) by
Seebach and co-workers,13 and their incorporation in
-peptide
b b
3- and 2-amino acid ho-
* Corresponding author. Tel.: þ33 540 003 020; fax: þ33 540 002 215; e-mail
address: g.guichard@iecb.u-bordeaux.fr (G. Guichard).
y
b
These authors contributed equally to the work.
0040-4020/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2011.11.066

Y.-R. Nelli et al. / Tetrahedron 68 (2012) 4492e4500
4493
foldamers for metal binding studies is noteworthy in this regard.14
Similarly, peptoids (oligomers of N-alkyl glycines) containing thiol
and imidazole side chains at defined position have been designed
that fold into two-helix bundles upon zinc addition.15
2.1. Synthesis of N-Boc-protected His-type activated
monomers 2a and 2b
We initially investigated the synthesis of monoprotected di-
amine 1b from Boc- -His(Ts)-OH. Reduction of Boc- -His(Ts)-OH
Aliphatic oligoureas bearing proteinaceous side chains (A) are
a class of non-peptide helical foldamers with potential to mimic
bioactive peptide helices and interact with bio(macro)molecules
a
a
into its corresponding histidinol using conditions developed by
Rodriguez and co-workers24 afforded alcohol 3 in modest yield
(32%). Attempts to activate alcohol 3 as its mesylate 4a or tosylate
4b did not yield any isolable products. Moreover, the direct con-
version of alcohol 3 into azide 5 under Mitsunobu conditions using
hydrazoic acid25 or diphenyl phosphorazidate (DPPA)26 as an al-
ternative failed. Furthermore, Thompson’s procedure using DPPA/
1,8-diazabicycloundec-7-ene (DBU) in toluene was also un-
successful (Scheme 1).27
(Fig. 1).16 These aza analogues of 4-peptides form well-defined 2.5
g
helical structures stabilized by three-centred H-bonds in solution17
and solid state.18 Aliphatic oligoureas are accessible by stepwise
synthesis using suitably protected activated building blocks derived
from ethylene diamine precursors (Fig. 1)19,20 or by condensation of
activated oligomeric segments.18a We have previously reported the
synthesis and use of N-tert-butoxycarbonyl (Boc)19a and N-fluo-
renylmethoxy-carbonyl (Fmoc) monomers19b of type B with side
chains of Ala, Val, Leu, Pro, Phe, Tyr, Trp and Lys. N-Fmoc-protected
building blocks were initially developed to allow the rapid synthesis
of oligoureas on solid support (Rink-amide resin).17,19b,21 Later, we
found that monomers protected with a Boc group were much more
effective, consistently giving oligoureas in higher yields and purities
following cleavage from the resin (4-methylbenzhydrylamine
(MBHA) resin).22 With the aim to expand our arsenal of proteino-
genic building blocks and to append histidine-type side chains at the
surface of oligourea helices, we have now investigated the synthesis
of a requisite N-Boc-protected building block starting from histidine.
Because of the nucleophilic character of the 1H-imidazole ring,
a judicious combination of N(a) and N(s) protecting groups on his-
Scheme 1. Synthetic strategy first envisioned for the preparation of azide 5. (i) MsCl or
tidine turned out to be crucial for the successful formation of His-
type monomers 2 and subsequent use for solid phase oligourea
synthesis.
TsCl, Et3N, CH2Cl2, 0 ꢀC; (ii) DIAD, PPh3, HN3, THF, 0 ꢀC, (iii) DPPA, DBU, NaN3, toluene.
At this point, as previously suggested by Wyatt in his work on
homologation of histidine,23 we suspected that the nucleophilicity
of the Boc-protected
a-amine was sufficient to cause an intra-
molecular attack of the activated alcohols 4 resulting in the deg-
radation of the reaction intermediates. In order to suppress the
reactivity of the histidine a-amino function, we decided to tem-
porarily convert it into a phthalimide group (Scheme 2).20b Sur-
prisingly, attempts to form the alcohol by reduction of the acid
function when using a N(a)-phthaloyl, N(s)-Ts protection combi-
nation did not yield any isolable product. This reduction was only
achieved when a trityl group (Trt) was installed on s-nitrogen and
under the conditions of McGeary28 to furnish alcohol 7 with
a moderate yield of 46% (Scheme 2). Several routes to convert 7 into
azide 9 were next investigated, but the formation of azide 9 was
only achieved by reaction of mesylate 8 with sodium azide, albeit in
very low yield (9%).
Fig. 1. Schematic representation of aliphatic oligoureas of type A and corresponding
monomeric B-type units. Formulae of targeted His-type activated monomers 2a and 2b
and corresponding N-Boc-protected ethylene diamine precursors 1a and 1b.
2. Results and discussion
Succinimidyl
{2-{[(tert-butoxy)carbonyl]amino}-2-X-ethyl}-
carbamate building blocks B bearing proteinogenic side chains have
been prepared either via a Curtius rearrangement of their corre-
sponding N-protected b
3-amino acids and subsequent trapping of
the isocyanate with N-hydroxy succinimide19 or by activation of the
corresponding N-protected ethylene diamine derivatives with N,N0-
disuccinimidyl carbonate (DSC).18a Several syntheses of N-Boc-
protected derivatives of
cluding Boc-
3hHis(Trt)-OH via the ArndteEistert homologation of
Boc-His(Tos)-OH by Seebach and co-workers13 and Boc- 3hHis(-
b
3-homohistidine have been reported, in-
b
b
Boc)-OH via the Kolbe nitrile synthesis of the beta amino alcohol
derived from Mts-His(Mts)-OH by Wyatt and co-workers.23 Al-
though these derivatives could eventually serve as precursors in the
synthesis of building block 2, the approach involving the prepara-
tion of a Boc-protected ethylene diamine precursor 1 was preferred
(Fig. 1).
Scheme 2. (i) (a) N-Carbethoxyphthalimide, Na2CO3; (b) Trt-Cl, Et3N, 83% over the two
steps. (ii) (a) BOP, DIEA, DMF, 0 ꢀC, (b) NaBH4, 46%; (iii) MsCl, Et3N, DCM, 0 ꢀC, 100%;
(iv) NaN3, DMF, 80 ꢀC, 9%. Inset: X-ray crystal structure of compound 7 (CCDC 844330).
See Supplementary data for details.

4494
Y.-R. Nelli et al. / Tetrahedron 68 (2012) 4492e4500
At this stage, we decided to reconsider our protecting group
strategy to access ethylene diamine 1 and we envisioned the in-
troduction of a temporary N( )-Trt protecting group on histidine.
The N( )-Trt group, which creates strong steric hindrance, was
expected to prevent intramolecular nucleophilic attack of any ac-
tivated intermediate. Previous studies with ditrityl histidine and
related ditrityl histamine derivatives have shown that the amino-
trityl is much more acid labile than the imino-trityl and that se-
lective N-detritylation is possible.29 Accordingly, we thus decided
version 14 by treatment with HCl and subsequent reaction with
Boc2O and DMAP (Scheme 4). Then, the N( )-Boc protection was
s
a
selectively removed by basic solvolysis with aqueous K2CO3 in
MeOH to give monoprotected derivative 15.32,33 Tosylation of 15
under standard conditions furnished key azide 5 in quantitative
yield. The structure of 5 was unambiguously characterized by X-ray
diffraction analysis (CCDC 844331) (Scheme 4). It is worth men-
tioning that 5, which was not accessible from Boc-His(Tos)-OH
(Scheme 1), was here obtained in good overall yield. Interestingly,
a
to keep the N(
s)-Trt protection on the imidazole ring and exploit
these results highlight the practical utility of the temporary N(a)-
the possibility for a selective N-Trt deprotection following azide
formation. Thus, 2HCl$H-His-OMe was treated with Trt-Cl in the
presence of triethylamine (Et3N) to afford Trt-His(s-Trt)-OMe 10 in
Trt protection to prevent nucleophilic intramolecular attacks and
other side reactions on activated intermediates as postulated
above.23 Azide 5 was next reduced to amine 1b by catalytic hy-
drogenation using 5% Pd/BaSO4 and 1b was finally converted to the
corresponding succinimidyl carbamate 2b in 88% yield over the last
two steps (Scheme 4).
quantitative yield (Scheme 3).30 Reduction of the methyl ester with
LiAlH4 furnished alcohol 11 in high yield.31 Several conditions for
the transformation of alcohol 11 into the corresponding azide were
screened. It was gratifying to find that in the presence of DPPA and
NaN3 alcohol 11 was converted to azide 12 in 75% yield. As antici-
pated, deprotection of 12 at the primary amine function occurred
selectively upon exposure to a 2% TFA solution in a mixture of
MeOH/DCE. The resulting TFA salt was immediately treated with
(Boc)2O in the presence of Et3N to furnish N-Boc-protected 13 in
good yield. Reduction of the azide function of 13 by hydrogenation
in presence of 5% Pd/BaSO4 afforded the N-Boc-protected ethylene
diamine 1a, which was transformed without further purification to
succinimidyl carbamate 2a after treatment with DSC in 68% over
the two steps (Scheme 3).
Scheme 4. (i) (a) Aqueous 6 M HCl, THF, 70 ꢀC, (b) (Boc)2O, Et3N, DMAP, CH3CN/H2O,
0
ꢀC, 65%; (ii) K2CO3, MeOH/H2O, 98%; (iii) TsCl, Et3N, DMAP, DCM, 0 ꢀC, 92%; (iv) H2,
Pd/BaSO4, EtOAc; (v) DSC, CH2Cl2, 86% over the two steps. Inset: X-ray crystal structure
of key azide 5. See Supplementary data for details.
In this second synthetic route featuring a selective N(s)-Boc
deprotection, His-derived carbamate 2b was obtained with an
overall yield of 51% starting from ditrityl azide 12 and in 33% from
commercially available 2HCl$H-His-OMe. Consequently, the origi-
nal strategy developed for the preparation of monomer 2a starting
from Trt-His(s-Trt)-OMe was successfully extended to access
Scheme 3. (i) Trt-Cl, Et3N, CH3CN; (ii) LiALH4, THF, 0 ꢀC, 93% over the two steps; (iii)
DPPA, DBU, NaN3, DMF, 75%; (iv) (a) DCE, MeOH, TFA, (b) (Boc)2O, Et3N, 76%; (v) H2, Pd/
BaSO4, EtOAc; (vi) DSC, CH2Cl2, 68% over the two steps.
monomer 2b. The utility of monomers 2a and 2b for the synthesis
on solid support of His-side chain-containing oligoureas was then
tested.
In this way, the synthesis of the N-Boc-protected succinimidyl
carbamate 2a equipped with a N(
was achieved in seven steps with an overall yield of 36%, starting
s
)-Trt protection on imidazole ring
2.2. Solid phase synthesis of oligoureas containing histidine-
type units
from commercially available 2HCl$H-His-OMe (Scheme 3).
However, the sensitivity of the N(
s
)-Trt group under the acidic
To enable comparison of building blocks 2a and 2b and facilitate
the analysis of the resulting imidazole-containing oligoureas,
a model sequence containing only one His-like unit was prepared.
Oligourea 16 was designed based on an antimicrobial amphiphilic
oligourea sequence21,22 by introduction of a His-type unit at the
penultimate position (Fig. 2). The general procedure for oligourea
synthesis on solid support is outlined in Scheme 5. The assembly of
the chain was performed manually starting from an MBHA resin
conditions required for Boc deprotection during chain extension, is
likely to be a limitation to the use of monomer 2a for oligourea
synthesis. For this reason, we explored the possibility to convert
azide precursor 12 into the potentially more useful monomer 2b
bearing a N(
s
)-Ts protection. By analogy to Boc-His(
s
-Tos)-OH,
)-Ts pro-
which is commonly used in peptide synthesis, the N(
s
tection in 2b is expected to be fully stable under the coupling and
Boc deprotection conditions during oligourea main chain elonga-
tion and to be removed during final HF cleavage of the resin.
Ditrityl-compound 12 was first converted into its diBoc-protected
(100 mmol scale) by sequential coupling of activated N-Boc-pro-
tected monomers B (3 equiv, 2ꢁ90 min) in the presence of diiso-
propylethylamine (DIEA) following a reported procedure (Scheme

Y.-R. Nelli et al. / Tetrahedron 68 (2012) 4492e4500
4495
5).22 Completion of the coupling steps was monitored by a chloranil
test,34 which compared to the Kaiser test35 proved to be more ac-
curate in determining the efficiency of the acylation steps.
was shown previously that this isosteric replacement at the ter-
minus has no consequence on the helix forming propensity of the
resulting urea oligomer.
Fig. 2. Imidazole-containing oligoureas 16e18 synthesized by SPS. Residue numbering starts from g
4-Val residue.
Whereas elongation of the chain starting from the g
4-Val resi-
due up to the second Trp-like residue proceeded smoothly, the
condensation of His-type monomer 2a, turned out to be more
difficult than expected. The chloranil test indicated incomplete
acylation even after prolonged coupling time. This difficulty may be
attributable to the bulkiness of the Trt group at the
imidazole ring, although the related trityl-protected histidine de-
rivative Fmoc-His( -Trt)-OH is commonly used in solid phase
s-position of the
s
peptide synthesis (SPPS). Alternatively, we cannot exclude that
helical folding of the oligourea during chain elongation may also
diminish the efficiency of this coupling step.37,38 Following Boc
deprotection and simultaneous removal of the N(s)-Trt protection
upon TFA treatment, the last residue (Val-type) and the terminal
capping motif (isopropylurea) were successively incorporated in
the growing chain and the resin was finally cleaved with HF. Re-
verse phase HPLC (RP-HPLC) coupled to electrospray ionization
mass spectrometry (ESI-MS) of the cleaved product indicated the
presence of two main compounds in a 3:2 ratio: the first one cor-
responding to the deletion oligomer missing the His-type unit, the
second one, with a molecular mass of 86 Da higher than the desired
oligomer 16, resulting from acylation of the imidazole group by
isopropylisocyanate during the last step of the SPS (17, Fig. 2). In
contrast, over-acylation of the imidazole side chain by the Val-type
monomer was not observed, suggesting that succinimidyl carba-
mates B are not reactive enough to react with the free imidazole
group. It is noteworthy that in aqueous acidic medium, purified
Scheme 5. General procedure for SPS of oligoureas. (i) Boc-g
4-Val-OH, BOP, DIEA, DMF.
(ii) Removal of the Boc protection: TFA, 2ꢁ5 min (iii) monomer B, DIEA, DMF, 2ꢁ3 h
(iv) (a) TFA, 2ꢁ5 min; (b) isopropylisocyanate, DIEA, DMF. (v) HF, 10% p-cresol, 0 ꢀC, 1 h.
The benzhydrylamine (BHA) and MBHA resins have been orig-
inally developed for the synthesis of amidated peptides using the
Boc strategy.36 However, the urea linkage formed by coupling
a Boc-protected monomer of type B to the amino group of an MBHA
or BHA resin is more acid labile than the corresponding amide
linkage and is not sufficiently stable to resist the acid treatment of
subsequent Boc deprotection rounds.22 To solve this problem and
prevent unwanted cleavage of the urea bond connected to the resin,
an amide bond is introduced first by coupling a N-Boc-protected g4
amino acid to the amino group of the MBHA resin (Scheme 5). It
-

4496
Y.-R. Nelli et al. / Tetrahedron 68 (2012) 4492e4500
oligomer 17 was found to undergo partial deacylation at the im-
idazole ring to give the target oligourea 16 (see Supplementary data
for details). Nevertheless, these observations underscore that un-
protected imidazole side chains in oligoureas are not compatible
with the use of isocyanates for the final capping step. The synthesis
of an oligomer uncapped at its terminus was thus undertaken.
Oligourea 18 (see Fig. 2) was obtained after HF cleavage, again as
a mixture with its deleted form (i.e., missing the His-type unit), in
a 1:1 ratio and was finally isolated after purification by preparative
RP-HPLC in 9% yield.
The NH/CH fingerprint region of the TOCSY spectrum of 16 is
shown in Fig. 3 and pairs of main chain diastereotopic CH2 pro-
tons are indicated for all residues. We previously reported that
the chemical shift difference (Dd) between main chain diaster-
eotopic CH2 protons is a useful descriptor of the conformational
homogeneity of helical N,N0-linked oligoureas.17,22,37 The diaster-
eotopicity values measured for 16 and 18 are collected in Fig. 4. In
both cases, Dd values for residues 2e6 are equal or larger than
1 ppm, which is consistent with a helical folding. However, the
amino terminal residue (P8) and the penultimate His-type resi-
due (P7) in 18 are characterized by no and weak diastereotopicity
of their methylene protons, respectively, which suggests helix
fraying in this part of the structure. In agreement with previous
studies on oligoureas,17,37 capping the amino group with iso-
propylisocyanate was found to reinforce the 2.5 helical pop-
ulation as evidenced by the significant increase of Dd values for
the last two residues of 16.
Finally, the solid phase synthesis of N-capped oligourea 16 was
repeated using monomer 2b. In contrast to what was observed with
2a, on resin urea bond formation with 2b was found to proceed to
completion after the second coupling step (based on chloranil test).
Thus, the benefits of a N(s)-Ts protection are twofold: first, it is less
hindered and as a result, promotes a better insertion of His-type
units within the oligourea backbone, and second, it remains in
place under Boc deprotection conditions. After Boc removal, and
introduction of the last residue, the chain was terminated by
treatment of the resin with isopropylisocyanate and the resin was
cleaved by HF. RP-HPLC analysis of the crude product revealed the
formation of a major compound (46% purity) whose identity was
confirmed by ESI-MS to be 8-mer 16 (see Supplementary data for
details). Oligourea 16 was purified by RP-HPLC and recovered in an
overall yield of 13% based on resin loading. The homogeneity of
oligoureas 16 and 18 was finally assessed by C18 RP-HPLC (>99%)
and their structures were confirmed by ESI-MS and 1H NMR anal-
yses. All together, these results demonstrate the superiority of
monomer 2b over 2a and its practical utility for the preparation of
oligourea foldamers containing His-type units.
2.3. 1H NMR conformational studies
NMR analysis was used to determine the helix forming pro-
pensities of oligoureas 16 and 18 in solution. 1H NMR spectra were
recorded in CD3OH at a concentration of 6 mM. Spin systems were
unambiguously resolved using a combination of COSY and TOCSY
2D experiments (for nomenclature of the main chain protons, see
formula A in Fig. 1).17 Sequence-specific assignment of all reso-
nances in the 1H NMR was achieved by a ROESY experiment and
was deduced from the strong N0H(iþ1),NH(i) NOE connectivities
within each urea bond. The helical character of 16 and 18 was
qualitatively inferred from the large vicinal coupling constants
Fig. 4. Chemical shift difference (Dd) between geminal aCH protons in oligoureas 16
(black bars) and 18 (grey bars) for residues 1e8 as determined by 1H NMR in
CD3OH.
2.4. Circular dichroism (CD) study
Circular dichroism is a useful and complementary technique to
study the folding propensity of aliphatic oligoureas. Far-UV CD
spectra of oligomers 16 and 18 were first recorded in tri-
fluoroethanol (TFE) at a concentration of 0.2 mM. As expected, CD
measurements revealed that in TFE, 16 retains a stable 2.5 helical
structure with a typical signature presenting an intense maximum
at 203 nm. The per residue molar ellipticity (PRME) value is com-
parable to that previously found for oligoureas of similar
length.17a,22 In contrast, the slight change observed in the shape of
the CD spectrum of uncapped 18 together with the significant de-
crease in the intensity of the CD maximum confirmed helix de-
stabilization observed by NMR (Fig. 5a). Overall, these results
emphasize the critical role played by the capping motif in stabi-
lizing 2.5 helical oligoureas in solution.
between NHs and
between vicinal coupling constants of main chain diastereotopic
methylene protons ( CH2) (Fig. 3 and Supplementary data).
bCH(R) protons, and the strong differentiation
a
Next, to gain information about the influence of the imidazole
side chain protonation state on the helicity of 16, we recorded CD
spectra in aqueous buffers within pH range from 5 to 7.6 (Fig. 5). It is
noteworthy that the CD signature characteristic of the canonical 2.5
helical structure observed in TFE is maintained in aqueous envi-
ronment at pH 7. However, the significantly lower PRME value at
l
¼203 nm indicates partial destabilization relative to TFE.39 Fur-
Fig. 3. Fingerprint NH/CH and N0H/CH region of the TOCSY experiment of oligomer 16,
recorded in CD3OH (400 MHz). For clarity reasons, a three letter code analogous to that
thermore, the CD signature was hardly affected by pH variations
between 5 and 7.6 thus suggesting that the protonation state of the
imidazole side chain has no influence on 2.5 helix folding pro-
pensity in water (Fig. 5b).
of standard a-amino acids with a U letter in superscript is used to identify all residues
in the sequence. Red and green dashed lines highlight the fingerprints of g
4-Val res-
idue and isopropyl group, respectively.

Y.-R. Nelli et al. / Tetrahedron 68 (2012) 4492e4500
4497
purchased from PolyPeptide Laboratories France. Benzotriazole-1-
yl-oxy-tris-(dimethylamino)-phosphonium hexa-fluorophosphate
(BOP) was purchased from Iris company. All organic solvents
were of analytical quality and Milli-Q (Millipore) water was used
for RP-HPLC analyses and purifications. Activated monomers used
for solid phase synthesis were prepared using a previously reported
procedure.21 Oligourea synthesis was performed manually in a sy-
ringe (vide infra). Thin layer chromatography (TLC) was performed
on silica gel 60 F254 (Merck) with detection by UV light and charring
with 1% ninhydrin in ethanol followed by heating. Flash column
chromatography was carried out on silica gel (40e63 mm, Merck).
Melting points were measured in open capillary tubes and are
uncorrected. Optical rotations were determined on a Jasco P-2000
polarimeter and are given as ½a D25
ꢂ
(concentration in g/100 mL sol-
vent). IR spectra were recorded with a Bruker IFS55 FT-IR spec-
trometer. 1H NMR and 13C NMR spectra were recorded on two
different NMR spectrometers: (1) an Avance II NMR spectrometer
(Bruker Biospin) with a vertical 7.05 T narrow-bore/ultrashield
magnet operating at 300 MHz for 1H observation and 75 MHz for
13C observation by means of a 5-mm direct BBO 1H/19F_XBB_H
probe with Z gradient capabilities; (2) a DPX-400 NMR spectrom-
eter (Bruker Biospin) with a vertical 9.4 T narrow-bore/ultrashield
magnet operating at 400 MHz for 1H observation by means of
a 5-mm direct QNP 1H/13C/31P/19F probe with gradient capabilities.
Chemical shifts are reported in parts per million (ppm) relative to
the 1H residual signal of the deuterated solvent used. 1H NMR
splitting patterns with observed first-order coupling are designated
as singlet (s), doublet (d), triplet (t), or quartet (q). Coupling con-
stants (J) are reported in hertz. CD spectra were recorded on a J-815
Jasco CD spectrometer. RP-HPLC analyses were performed on
a Dionex U3000SD using a MachereyeNagel Nucleodur column
(4.6ꢁ100 mm, 3
m
m) at a flow rate of 1 mL minꢃ1. The mobile phase
was composed of 0.1% (v/v) TFA/H2O (Solvent A) and 0.1%TFA/
CH3CN (Solvent B). Semi-preparative purifications of oligoureas
were performed on a Gilson GX-281 system using a Macher-
Fig. 5. (a) Far-UV CD spectra of 16 and 18 recorded at 0.2 mM in TFE. (b) Far-UV CD
spectra of 16 recorded at 0.2 mM in aqueous buffers at different pH values (10 mM
sodium acetate and phosphate buffers). Data are expressed in term of molar ellipticity
per residue ([q
] in deg cm2 dmolꢃ1).
eyeNagel Nucleodur column (20ꢁ250 mm, 5
mm) at a flow rate of
20 mL minꢃ1. The mobile phase was similar as for the analytic
system, unless otherwise notified. A gradient elution (0e20 min:
80%e40% A) was applied at a flow rate of 20 mL minꢃ1. Column
effluent was monitored by UV detection at 214 and 254 nm. ESI-MS
analyses were carried out on a ThermoElectron LCQ Advantage
spectrometer equipped with an ion trap mass analyzer and coupled
with a ThermoElectron Surveyor HPLC system.
3. Conclusion
An efficient synthesis of activated succinimidyl carbamate of
type B (PG¼Boc) with (1H-imidazol-4-yl)-methyl side chain (2a and
2b) has been developed starting from histidine. After several un-
successful attempts, we found that the protecting group scheme on
histidine was critical to access key monoprotected diamine pre-
cursors (1a and 1b). Only when starting from Trt-His(s-Trt)-OMe,
4.2. Synthesis of building blocks 2a and 2b
could N-Boc-protected diamines 1 be obtained in satisfactory yields.
Activated building blocks 2a and 2b were finally obtained in 36% and
33% overall yield, respectively, starting from commercially available
2HCl$H-His-OMe. The utility of 2b and its superiority over 2a for the
SPS of aliphatic oligourea foldamers incorporating His-type units
has been demonstrated. The preparation of 8-mer oligourea 16
containing one imidazole side chain was successfully achieved in
good overall yield and purity when employing 2b. Conformational
analyses by 1H NMR spectroscopy and CD confirmed helical folding
of 16 in both TFE and aqueous environment. Access to new oligourea
sequences with His-type blocks at selected positions is likely to be
useful for the design of more sophisticated foldamer architectures
and will certainly contribute to expand the scope of their possible
applications (e.g., metal ion recognition and/or catalysis).
4.2.1. (S)-Methyl 3-(1-trityl-1H-imidazol-4-yl)-2-(tritylamino)prop-
anoate (10). To a stirred suspension of 2HCl$H-His-OMe (15 g,
61.98 mmol) in CH3CN (600 mL) at
309.92 mmol) was added and after
0
5
ꢀC, Et3N (43.31 ml,
min Trt-Cl (43.07 g,
154.96 mmol) was added slowly portion wise. The reaction mixture
was allowed to stir at room temperature overnight. The CH3CN was
concentrated under reduced pressure and the residue was dis-
solved in CH2Cl2 (500 mL), washed with aqueous saturated solution
of NaHCO3 (100 mL), water (100 mL) and with brine (100 mL), dried
over MgSO4 and concentrated under high vacuum to afford 10
(40.47 g, quantitative yield) as a yellow solid, which was used for
next step without characterization.
4.2.2. (S)-3-(1-Trityl-1H-imidazol-4-yl)-2-(tritylamino)propan-1-ol
(11). To a stirred suspension of LiAlH4 (9.39 g, 247.90 mmol) in dry
THF (100 ml) at 0 ꢀC, compound 10 (40.47 g, 61.97 mmol) dissolved
in dry THF (300 ml) was added slowly and the reaction mixture was
allowed to stir a room temperature for 2 h. The reaction mixture
was carefully quenched with aqueous saturated solution of Na2SO4,
diluted with EtOAc (400 mL), and filtered over Celite bed. The
4. Experimental section
4.1. General methods
Commercially available reagents were used throughout without
purification. Boc-g
4-Val-OH and MBHA resin (1 mmol/g) were

4498
Y.-R. Nelli et al. / Tetrahedron 68 (2012) 4492e4500
filtrate was washed with water (100 mL), brine (100 mL), dried over
Na2SO4, and concentrated under vacuum. After purification on
silica gel eluting with 5% EtOAc in cyclohexane to EtOAc/CH2Cl2
(1:1, v/v), alcohol 11 was isolated as a solid (36.1 g, 93%). 1H NMR
temperature for 3 h. The reaction mixture was then filtered off, the
filtrate was washed with an aqueous molar solution of KHSO4
(3ꢁ5 mL), the organic layer was dried over MgSO4 and concen-
trated under vacuum, and 2a was obtained after recrystallization
using CH2Cl2/petroleum ether solvent mixture (3.21 g, 68%). Mp
124e127 ꢀC; IR (neat): 3402,1770, 1749, 1730, 1501, 1481, 1178, 1163,
(300 MHz, CDCl3):
d
(ppm) 2.00 (d, J¼6.1, 14.4 Hz, 2H), 2.45 (d,
J¼14.4 Hz, 1H), 2.88e3.14 (m, 2H), 3.55 (d, J¼11.1 Hz, 1H), 6.35 (s,
1H), 7.08e7.44 (m, 25H), 7.55 (d, J¼6.9 Hz, 6H); 13C NMR (75 MHz,
1145 cmꢃ1 1H NMR (300 MHz, CDCl3):
; d 1.31 (s, 9H), 2.75 (s, 4H),
CDCl3):
d
(ppm) 31.8, 52.5, 65.8, 71.0, 75.2, 120.4, 126.2, 127.8, 128.1,
3.23e3.02 (m, 2H), 3.48e3.39 (m, 2H), 4.07e3.92 (m, 1H), 6.24 (d,
J¼8.0 Hz, 1H), 6.79 (s, 1H), 7.20e7.06 (m, 6H), 7.50e7.34 (m, 9H),
128.7, 129.7, 137.7, 138.1, 142.3, 147.2.
7.75e7.64 (m, 1H), 8.21 (s, 1H); 13C NMR (75 MHz, CDCl3):
d 25.5,
4.2.3. (S)-1-Azido-N-trityl-3-(1-trityl-1H-imidazol-4-yl)propan-2-
amine (12). To a stirred solution of alcohol 11 (36.1 g, 55.54 mmol)
in DMF (180 ml) at 65 ꢀC, were successively added DPPA (17.85 mL,
83.30 mmol), DBU (11.74 mL, 83.30 mmol) and after 5 h NaN3
(1.81 g, 27.769 mmol). Stirring was maintained at the same tem-
perature for 12 h after which time the reaction mixture was cooled
down to room temperature, then diluted with EtOAc (360 mL) and
washed with saturated solution of NaHCO3 (100 mL), water
(2ꢁ100 mL), brine (100 mL), dried over Na2SO4, and concentrated
under reduced pressure. After purification on silica gel eluting with
6% EtOAc in cyclohexane azide 12 was obtained as an off white solid
27.1, 28.3, 44.7, 50.3, 78.2, 79.1, 120.7, 128.6, 128.9, 129.8, 132.5,
135.7, 140.3, 152.8, 155.9, 170.4; HRMS(ESI): m/z calculated for
C35H38N5O6 [MþH]þ, 624.2822 found 624.2819.
4.2.7. (S)-tert-Butyl 4-(3-azido-2-(tert-butoxycarbonylamino)-pro-
pyl)-1H-imidazole-1-carboxylate (14). To a stirred solution of azide
12 (2.28 g, 3.507 mmol) in THF (10 mL) at room temperature, a so-
lution of aqueous 6 N HCl (10 mL) was added and stirring was
maintained at 70 ꢀC for 4 h. After completion of the reaction, the
THF was removed under reduced pressure, and water (10 ml) was
added. The aqueous phase was washed with CH2Cl2 (2ꢁ5 mL). The
aqueous layer was concentrated then co-evaporated with EtOH
under reduced pressure and dried under high vacuum line to fur-
nish hydrochloride salt, which was directly suspended in CH3CN
(20 mL) at 0 ꢀC. Et3N (0.487 mL, 3.51 mmol), DMAP (0.042 g,
0.35 mmol) followed by Boc2O (1.77 mL, 7.72 mmol) and of water
(0.5 mL) were successively added and the reaction mixture was
allowed to stir at room temperature overnight. Acetonitrile was
concentrated under vacuum and the crude was dissolved in EtOAc
(40 mL), the organic layer was washed with water (5 mL), brine
(5 mL), and dried with MgSO4 and concentrated under reduced
pressure. After silica gel purification eluting with 15% EtOAc in
cyclohexane compound 14 was obtained (0.832 g, 65%) as a white
a 25
(28.1 g, 75%). Mp 67e69 C; ½ ꢂD þ11.6 (c 1.0, CHCl3); IR (neat):
ꢀ
3030, 2098, 1490, 1446, 1185, 1130, 1056, 1033, 902 cmꢃ1 1H NMR
(300 MHz, CDCl3):
;
d
2.22 (dd, J¼7.2, 14.4 Hz, 1H) 2.37 (d, J¼7.3 Hz,
1H), 2.48 (dd, J¼4.6, 14.4 Hz, 1H), 2.71 (dd, J¼6.3, 11.8 Hz, 1H),
3.01e2.89 (m, 1H), 3.08 (dd, J¼3.3, 11.8 Hz, 1H), 6.41 (d, J¼1.2 Hz,
1H), 7.43e7.04 (m, 25H), 7.58e7.49 (m, 6H); 13C NMR (75 MHz,
CDCl3):
d 31.2, 52.2, 54.3, 71.1, 75.2, 77.2, 120.0, 125.6, 127.9, 128.0,
128.7, 129.8 137.7, 138.4, 142.3, 146.7; HRMS(ESI):m/z calculated for
C44H38N6 [MþH]þ 651.3236, found 651.3242.
4.2.4. (S)-tert-Butyl 1-azido-3-(1-trityl-1H-imidazol-4-yl)propan-2-
ylcarbamate (13). Azide 12 (6.1 g, 9.38 mmol) was dissolved in
a C2H4Cl2/MeOH/TFA (188:8:4, v/v/v) solution mixture (200 mL).
The reaction mixture was stirred between 0 and 10 ꢀC for 10 min,
then basified with Et3N, then treated with (Boc)2O (2.15 ml,
9.38 mmol) and allowed to stir at room temperature for 20 min.
The reaction mixture was washed with saturated aqueous solution
of NaHCO3 (50 mL), with molar aqueous solution of KHSO4 (50 mL),
water (50 mL) and with brine (50 mL), dried over MgSO4, and
concentrated under vacuum., After purification on silica gel eluting
with 10e20% of EtOAc in cyclohexane, azide 13 was isolated as an
solid. Mp 81e83 ꢀC; ½a D25
þ2.4 (c 1.0, CHCl3); IR (neat): 3421, 2951,
ꢂ
2092, 1748, 1720, 1532, 1505, 1396, 1384, 1261, 1250, 1164 cmꢃ1, 1H
NMR (300 MHz, CDCl3):
d
1.31 (s, 9H) 1.50 (s, 9H), 2.70 (d, J¼6.0 Hz,
2H), 3.30 (dd, J¼5.1, 12.1 Hz, 1H), 3.33 (dd, J¼5.1, 12.1 Hz, 1H),
3.98e3.82 (m, 1H), 5.52 (d, J¼7.5 Hz, 1H), 7.09 (s, 1H), 7.91 (d,
J¼1.1 Hz, 1H); 13C NMR (75 MHz, CDCl3):
d 27.7, 28.2, 29.8, 49.9,
53.2, 79.2, 85.4, 114.6, 136.7, 139.1, 146.9, 155.0; HRMS(ESI): m/z
calculated for C16H27N6O4 [MþH]þ, 367.2094 found 367.2093.
off white solid (3.65 g, 76%). Mp 47e50 ꢀC; ½a D25
ꢂ
ꢃ6.9 (c 1.0, CHCl3);
4.2.8. (S)-tert-Butyl-1-azido-3-(1H-imidazol-4-yl)propan-2-
ylcarbamate (15). To a stirred solution of compound 14 (0.400 g,
1.09 mmol) in MeOH (4 mL) at 0 ꢀC, K2CO3 (0.075 g, 0.55 mmol) and
water (0.1 mL) were added and the reaction was allowed to stir at
room temperature for 1 h. MeOH was removed under reduced
pressure, and the crude was dissolved in EtOAc (20 mL). The or-
ganic layer was washed with water (5 mL), brine (5 mL), dried with
MgSO4 and concentrated under reduced pressure to afford after
drying under high vacuum line compound 15 (0.285 g, 98%) as an
IR (neat): 3025, 2099, 1706, 1494, 1446, 1365, 1242, 1167, 1036 cmꢃ1
.
1H NMR (300 MHz, CDCl3):
(dd, J¼6.8, 12.1 Hz, 1H), 3.48 (dd, J¼5.3, 12.1 Hz, 1H), 4.05e3.90 (m,
1H), 5.92 (d, 1H, J¼5.7 Hz), 6.74 (s, 1H), 7.18e7.08 (m, 6H), 7.44e7.32
d
1.41 (s, 9H), 2.75 (d, J¼5.7 Hz, 2H), 3.28
(m, 9H), 7.66 (br s, 1H); 13C NMR (75 MHz, CDCl3):
d 26.9, 28.3 (3),
29.3, 50.2, 53.4, 75.9, 79.3, 120.1, 128.2 (2), 128.3 (4),129.6 (4), 135.9,
137.9, 141.8 (3), 155.4; HRMS(ESI): m/z calculated for C30H33N6O2
[MþH]þ 509.2665, found 509.2657.
off white solid. Mp 86e87 ꢀC; ½a D25
ꢃ9.5 (c 1.0, CHCl3); IR (neat):
ꢂ
4.2.5. (S)-tert-Butyl 1-amino-3-(1-trityl-1H-imidazol-4-yl)propan-
2-ylcarbamate (1a). To a stirred solution of 13 (3.61 g, 0.39 mmol)
in EtOAc (72 mL) at room temperature, 5% Pd on BaSO4 (400 mg)
was added and the reaction mixture was stirred under H2 gas at-
mosphere for 20 h. The reaction mixture was filtered over Millipore
filter paper, the filtrate was concentrated under reduced pressure
and dried under high vacuum line to afford amine 1a (3.41 g,
quantitative yield) as a pale yellow liquid, which was used for next
step without characterization.
3392, 2980, 2089, 1710, 1530, 1504, 1391, 1251, 1130 cmꢃ1, 1H NMR
(300 MHz, CDCl3):
d
1.42 (s, 9H), 2.86 (d, J¼6.3 Hz, 2H), 3.30 (dd,
J¼6.1, 12.1 Hz, 1H), 3.42 (dd, J¼4.9, 12.2 Hz, 1H), 4.10e3.90 (m, 1H),
5.56 (d, 1H), 6.87 (s, J¼8.5 Hz, 1H), 7.62 (d, J¼1.0 Hz, 1H), 10.33 (br s,
1H); 13C NMR (75 MHz, CDCl3):
d 28.3, 29.4, 50.2, 53.3, 79.8, 116.0,
134.0, 135.2, 155.5; HRMS(ESI): m/z calculated for C11H19N6O2
[MþH]þ, 267.1569 found 267.1571.
4.2.9. (S)-tert-Butyl-1-azido-3-(1-tosyl-1H-imidazol-4-yl)propan-2-
ylcarbamate (5). To a stirred solution of 15 (0.285 g, 1.071 mmol) in
CH2Cl2 (5 ml) at 0 ꢀC, Et3N (0.178 ml, 1.285 mmol), followed by tosyl
chloride (0.224 g, 1.18 mmol) and DMAP (0.013 g, 0.11 mmol) were
added and the reaction mixture was stirred at room temperature
for 12 h. The reaction mixture was then diluted with CH2Cl2
4.2.6. Carbamate (2a). To
a
stirred suspension of N,N0-dis-
uccinimidyl carbonate (2.174 g, 8.50 mmol) in CH2Cl2 (10 mL),
a solution of amine 1a in CH2Cl2 (20 mL) was added dropwise
(3.41 g, 7.07 mmol) and the reaction mixture was stirred at room

Y.-R. Nelli et al. / Tetrahedron 68 (2012) 4492e4500
4499
(15 mL), washed with molar aqueous solution of KHSO4 (5 mL),
water (5 mL), brine (5 mL) dried with MgSO4, and concentrated
under reduced pressure. After silica gel purification eluting with
15% EtOAc in cyclohexane compound 5 was obtained as white solid
(HOBt) in DMF for 15 min. Other side chain deprotections and
cleavage of the oligomers from the resin were performed simulta-
neously by treatment with HF (containing 10% p-cresol as a scav-
enger) for 60 min at 0 ꢀC. The crude oligoureas were finally purified
by RP-HPLC and lyophilized.
(0.412 g, 92%). Mp 104e106 ꢀC; ½a D25
ꢃ2.0 (c 1.0, CHCl3); IR (neat):
ꢂ
3438, 2096, 1709, 1512, 1392, 1287, 1260, 1141, 1089 cmꢃ1; 1H NMR
Oligomer 16: Purity of crude product 46%. Yield after purification
19 mg, 13%; white powder; RP-HPLC tR 7.71 min (linear gradient,
20e60% B, 10 min); MS (ESI) m/z 1453.7 [MþH]þ.
(300 MHz, CDCl3):
d
1.40 (s, 9H), 2.43 (s, 3H), 2.75 (d, J¼5.5 Hz, 2H),
3.25 (dd, 1H, J¼6.1, 12.2 Hz), 3.38 (dd, 1H, J¼4.8, 12.2 Hz), 4.05e3.89
(m, 1H), 5.30 (d, J¼6.8 Hz, 1H), 7.05 (d, J¼1.0 Hz, 1H), 7.35 (d,
J¼8.2 Hz, 2H), 7.83 (d, J¼8.5 Hz, 2H), 7.93 (d, 1H, J¼1.3 Hz); 13C NMR
Oligomer 17: Purity of crude product 31%. Yield after purification
3.5 mg, 5%; white powder; RP-HPLC tR 8.34 min (linear gradient,
20e60% B, 10 min); MS (ESI) m/z 1538.7 [MþH]þ.
(75 MHz, CDCl3):
d 21.7, 28.3, 29.9, 49.6, 53.2, 79.6, 114.8, 127.3,
130.8,134.8,136.4,140.7,146.3,155.1; HRMS(ESI): m/z calculated for
Oligomer 18: Purity of crude product 19%. Yield after purification
11 mg, 8%; white powder; RP-HPLC tR 5.21 min (linear gradient,
20e60% B, 10 min); MS (ESI) m/z 1367.5 [MþH]þ.
C18H25N6O4S [MþH]þ, 421.1658 found 421.1660.
4.2.10. (S)-tert-Butyl 1-amino-3-(1-tosyl-1H-imidazol-4-yl)propan-
2-ylcarbamate (1b). To a stirred solution of azide 5 (0.150 g,
0.36 mmol) in EtOAc (20 ml) at room temperature 5% Pd on BaSO4
(0.050 g) was added and the reaction was stirred under H2 gas at-
mosphere for 20 h. The reaction mixture was filtered over Millipore
filter paper, the filtrate was evaporated and dried under high vac-
uum to afford compound 1b (0.140 g, quantitative yield) as a pale
yellow liquid, which was directly used for next step without
characterization.
Acknowledgements
This work was supported in part by the Centre National de la
Recherche Scientifique (CNRS), the Region Aquitaine, the University
of Bordeaux, and by a grant from the Agence Nationale de la
Recherche (grant no. ANR-07-PCVI-0018). A CIFRE fellowship from
ImmuPharma France and the Association Nationale pour la
Recherche Technique (ANRT) for P.C. is gratefully acknowledged.
ꢀ
The authors thank Axelle Grelard for her assistance with NMR
experiments.
4.2.11. Carbamate (2b). To a stirred suspension of DSC (0.109 g,
0.43 mmol) in CH2Cl2 (5 mL), a solution of amine 1b (0.140 g,
0.355 mmol) in CH2Cl2 (5 mL) was added dropwise at 0 ꢀC and left
to react at room temperature for 3 h. The reaction mixture was
filtered off, the filtrate was washed with molar aqueous solution of
KHSO4 (2ꢁ5 mL). The organic layer was dried over MgSO4 and
concentrated under reduced pressure. Compound 2b was isolated
after recrystallization with a mixture of CH2Cl2 and petroleum
ether as a white solid (0.165 g, 86%). Mp 74e76 ꢀC; IR (neat): 3342,
1762, 1752, 1720, 1540, 1506, 1410, 1390, 1180, 1174, 1142,
Supplementary data
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.tet.2011.11.066. These data in-
clude MOL files and InChiKeys of the most important compounds
described in this article.
1091 cmꢃ1 1H NMR (300 MHz, CDCl3):
; d 1.40 (s, 9H), 2.45 (s, 3H),
References and notes
2.77 (d, J¼5.7 Hz, 2H), 2.82 (s, 4H), 3.39e3.17 (m, 2H), 4.06e3.88 (m,
1H), 5.48 (d, J¼7.2 Hz, 1H), 6.82 (br s, 1H), 7.30 (s, 1H), 7.38 (d,
J¼8.3 Hz, 2H), 7.83 (d, J¼8.3 Hz, 2H), 7.97 (d, J¼1.1 Hz, 1H); 13C NMR
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