In-Peptide Synthesis of Imidazolidin-2-one Scaffolds, Equippable with Proteinogenic or Taggable/Linkable Side Chains, General Promoters of Unusual Secondary Structures

Article abstract of DOI:10.1021/acs.joc.8b03055

Peptidomimetics containing (S)- or (R)-imidazolidin-2-one-4-carboxylate (Imi) have been obtained by the expedient in-peptide cyclization of (S)- or (R)-α,β-diaminopropionic acid (Dap) residues. These Imi scaffolds behave as proline analogues characterized by a flat structure and a trans-restricted geometry of the preceding peptide bond and induce well-defined secondary structures in a biomimetic environment. While (S)-Imi peptides adopted a γ′-turn conformation, (R)-Imi induced the contemporary formation of a γ-turn and a rare 11-membered H-bonded structure in the 2→4 opposite direction of the sequence, identified as a ?-turn. In order to exploit these Imi scaffolds as general promoters of unusual secondary structures, proteinaceous side chains have been introduced at the N₁ position of the five-membered ring, potentially mimicking any residues. Finally, the Imi rings have been equipped with unnatural side chains or with functionalized substituents, which can be utilized as linkers to chemoselectively bind the Imi-peptides onto nanoparticles, biomaterials, or diagnostic probes.

Full text of DOI:10.1021/acs.joc.8b03055

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Article
In-peptide synthesis of imidazolidin-2-one scaffolds,
equippable with proteinogenic or taggable/linkable side
chains, general promoters of unusual secondary structures
Rossella De Marco, Junwei Zhao, Arianna Greco, Simone Ioannone, and Luca Gentilucci
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.8b03055 • Publication Date (Web): 08 Apr 2019
Downloaded from http://pubs.acs.org on April 9, 2019
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In-peptide synthesis of imidazolidin-2-one scaffolds, equippable with
proteinogenic or taggable/linkable side chains, general promoters of unusual
secondary structures
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Rossella De Marco,§ Junwei Zhao, Arianna Greco, Simone Ioannone, and Luca Gentilucci *
Dept. of Chemistry “G. Ciamician”, University of Bologna, via Selmi 2, 40126 - Bologna, Italy
* E-mail: luca.gentilucci@unibo.it; Fax: +39 0512099456; Phone: +39 0512099570; ORCID: 0000-0001-
9134-3161
§ rossella.demarco2@unibo.it
Table of Contents/Abstract Graphic
Abstract: Peptidomimetics containing (S)- or (R)-imidazolidin-2-one-4-carboxylate (Imi) have
been obtained by the expedient in-peptide cyclization of (S)- or (R)-,-diaminopropionic acid
(Dap) residues. These Imi scaffolds behave as proline analogues characterized by a flat structure
and a trans-restricted geometry of the preceding peptide bond, and induce well-defined secondary
structures in a biomimetic environment. While (S)-Imi peptides adopted a '-turn conformation, (R)-
Imi induced the contemporary formation of a -turn and a rare 11-membered H-bonded structure in
the 2→4 opposite direction of the sequence, identified as a -turn. In order to exploit these Imi
scaffolds as general promoters of unusual secondary structures, proteinaceous side chains have been
introduced at the N1 position of the five membered ring, potentially mimicking any residues.
Finally, the Imi rings have been equipped with unnatural side chains, or with functionalized
substituents, which can be utilized as linkers to chemoselectively bind the Imi-peptides onto
nanoparticles, biomaterials, or diagnostic probes.
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Introduction
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Heterocyclic-based peptidomimetics have been widely utilized to increase metabolic and
conformational stability of the parent peptides.1 Amid them, N-heterocycles are prevalent in
biologically active peptides and are increasingly attractive scaffolds in the development of new
pharmaceuticals.2 Relevant examples include pseudo-prolines,3 Freidinger-Veber lactams or
analogues,4 and cyclic urea scaffolds.5
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In particular, the latter have been utilized as structural elements in peptidomimetic inhibitors
of HIV protease and HIV replication,6 antibacterial MurB inhibitors,7 dopamine D4 and CGRP
receptor antagonists,8 ACE inhibitors,9 serine protease inhibitors,10 integrin inhibitors.11 In
literature, cyclic ureas are most commonly constructed via treatment of 1,2-diamine precursors with
carbonyldiimidazole,6 or by intramolecular diamination of alkenes,12 via rearrangement of Asn,9,13
via ring expansion of aziridine derivatives,14 by cyclization of aza-propargylglycinamides,5b by
alkylation of the urea nitrogen of semicarbazone residues,15 by Pd-catalyzed urea carboamination
reactions,16 and so on. Although a number of synthetic methods are available, many are not feasible
in peptide chemistry. Thus, alternative strategies for the construction of cyclic ureas would provide
more facile access to a class of peptidomimetics that are not readily available.
In this context, herein we report the expedient synthesis of peptidomimetics containing
imidazolidin-2-one-4-carboxylate (Imi) scaffolds, by cyclization of sequences containing ,-
diaminopropionic acid (Dap). The unexpected conformational features of the Imi-peptides are also
discussed with the aid of NMR analysis and molecular dynamics simulations.
Furthermore, peptides containing N-substituted Dap have been obtained from -
iodoalanine, by displacement with N-nucleophiles. The cyclization of the substituted Dap residues
gave access to Imi-peptides carrying proteinogenic or unnatural side chains at the position 1, or
eventually providing useful handles at which to undertake site-selective modifications of the
sequences.17 As a proof of concept, the allyl substituent at the position 1 of the Imi ring was
selectively derivatized by the Heck reaction.
Results and Discussion
Imi-peptides synthesis. Model peptides 2 containing (S)- or (R)-Imi scaffolds have been obtained
by the easy cyclization of the corresponding sequences 1 containing Dap (Scheme 1 and Table 1).
The preparation of the peptides 1a-c was conducted by coupling in solution Boc-protected amino
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acids to amino ester counterparts under MW irradiation,18 using a microwave oven specifically
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6
.
designed for organic synthesis, with N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC) HCl
and 1-hydroxybenzotriazole (HOBt) as activating agents, in the presence of trimethylamine (TEA).
Several Dap protected variants are commercially available; herein, Dap was introduced as Boc-
Dap(Cbz)-OH. In any case, Dap can be rapidly prepared following the literature19 and protected
thereafter with the appropriate group. Boc deprotection was performed with 25 % TFA in DCM.
The intermediates, obtained in quantitative yield, were not purified, while the final sequences 1a-c
were isolated in 80-85 % yield by flash chromatography over silica-gel (eluent 1:2
cyclohexane/EtOAc). Their identity was confirmed by ESI MS analysis, and purity was assessed to
be 80-90 % by RP HPLC.
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After removal of the N-Cbz group by catalytic hydrogenation, the Dap-peptides were
treated without further purifications with N,N′-disuccinimidyl carbonate (DSC) 3b,c and DIPEA in
3:1 DCM/DMF, giving the Imi-peptides 2a-c in good yield (86-91 %) after isolation by semi-
preparative RP HPLC (Scheme 1, Table 1), > 90 % pure as determined by analytical RP HPLC. The
identity of the compounds 2a-c (and of the Imi-peptides described in the next sections) was
confirmed by exact mass, proton-decoupled 13C NMR, and 1H NMR (the unambiguous assignment
of the resonances was done by 2D gCOSY).
Cbz
NH
H
N
O
O
H
1. H2 Pd/C
N
N
R
Boc
R
Boc
N
N
H
2. DSC (1.2 eq)
DIPEA (1.2 eq)
H
O
O
O
1
86-91%
2
a
c
b
(S)-Dap, R = Phe-OMe; (S)-Dap, R = Phe-Val-OMe;
(R)-Dap, R = Phe-Val-OMe
Scheme 1. In-solution preparation of Imi-peptides 2 from Dap-peptides 1.
Table 1. Synthesis of model Imi-peptides 2 by cyclization of the corresponding Dap-containing
peptides with DSC/DIPEA.
2
a
b
c
sequence
Peptide synthesis
yield %
86b
purity %a
Boc-Ala-(S)-Imi-Phe-OMe
Boc-Ala-(S)-Imi-Phe-Val-OMe
Boc-Ala-(R)-Imi-Phe-Val-OMe
in-solution
92
93
93
“
“
88b
91b
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d
Boc-Val-Ala-(S)-Imi-Phe-OMe
SPPS
SPPS
84c
86c
95
96
e
Boc-Val-Ala-(R)-Imi-Phe-OMe
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8
a
b
c
Determined by RP HPLC. Yield of the cyclization step, after isolation by RP HPLC. Calculated on the
9
average resin load, after isolation by RP HPLC.
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Also, Imi-peptides were successfully obtained by solid phase peptide synthesis (SPPS)
(Scheme 2). The tetrapeptides Boc-Val-Ala-(S)/(R)-Dap(Cbz)-Phe-resin (1d, 1e) were prepared
onto a Phe-preloaded Wang resin using standard Fmoc-protected amino acids and the
.
EDC HCl/HOBt/TEA coupling agents, under MW irradiation. Fmoc was removed by treatment
with piperidine/DMF. After deprotection of the Cbz group by catalytic hydrogenation with
ammonium formate (4 equiv.) in isopropanol/toluene,20 cyclization was done on resin with DSC
and DIPEA (3 equiv. each) under recently refined reaction conditions.21 Subsequently, the cleavage
from the resin with refluxing MeOH/TEA (Scheme 2)22 gave the peptides 2d or 2e, isolated in a
very satisfactory yield and purity (84 and 86 %, respectively, calculated on the basis of the average
resin load, Table 1) as described above. These peptides were prepared as methyl esters and with
Boc at the N-terminus, for structural comparison with the peptides 2b,c (see next paragraph).
Cbz
HN
1. Pd/C
HCO2NH4 (4 eq),
MW
H
O
H
O
N
H
N
N
N
H
O
O
Boc
O
2. DSC (3 eq),
DIPEA (3 eq)
Ph
1
d
: (S)-Dap
: (R)-Dap
3. TEA/MeOH, 
e
H
N
O
N
O
d
e
2
: (S)-Imi 84%
: (R)-Imi 86%
H
O
H
N
N
N
H
O
OMe
Boc
O
Ph
Scheme 2. SPPS of Imi-peptides 2d or 2e.
Epimerization of Dap during peptide synthesis and cyclization, in solution or in solid phase,
was excluded on the basis of the analysis of the reaction mixtures of 2a-c by HPLC MS and NMR.
Conformational analysis. X-ray crystallographic analysis of 3-acyl-imidazolidine-2-one-4-
carboxylates showed that these heterocycles have a planar imidazolidine-2-one ring, and the amide
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bond at the 3-position is restricted to the trans geometry, coplanar with the five-membered ring.23
Theoretical computations quantified that the differences in potential energy between the cis- and
trans-conformers were so large that the unstable cis-conformer would have an extremely low
concentration.23 This is in contrast to proline, which shows a puckered conformation of the
pyrrolidine ring, and is involved in a trans/cis equilibrium about the preceding amide bond.24
Hence, it is expected that the introduction of Imi scaffolds into a peptide sequence might impose
alternative conformations and unusual secondary structures. Previous investigations on
oligopeptides containing oxazolidin-2-one-4-carboxylates as pseudo-prolines showed the ability of
the heterocycles to stabilize turns, foldamers,3c-d,25 or even to form hydrogels.26
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In this perspective, the conformational bias exerted by the Imi scaffold on the overall
structure of model tetrapeptides was analyzed by NMR experiments at 400 MHz in 8:2
[D6]DMSO/H2O, a solvent mixture recommended by several authors as an excellent biomimetic
medium, in which intermolecular interactions are generally negligible for peptide concentrations in
the millimolar range.27 In any case, self-association of the peptides was excluded based on the
absence of concentration effects on the chemical shift of non-exchangeable protons.25,28 As
representative examples, we analyzed the tetrapeptides 2b and 2c, which include (S)- or (R)-
configured Imi, respectively, at the position 2 of the sequence, and peptides 2d and 2e having (S)-
or (R)-Imi at the position 3. All spectra showed a single set of sharp resonances, suggestive of
conformational homogeneity or of a fast equilibrium between slightly different conformers
(Supporting Information). gCOSY experiments were utilized for the unambiguous assignment of
the resonances.
In analogy to what observed for peptides containing oxazolidin-2-one-4-carboxylates, a
nonconventional ImiC=O⋯H-C(-1) intramolecular H-bond was expected (Figure1).3c-d,25 Indeed,
1
the H NMR analyses of all of the compounds showed a significantly downfield position of the -
proton of the residues preceding the Imi rings, consistent with the deshielding effect exerted by
ImiC=O for a trans conformation of the amide bond between Ala1-Imi2 (Figure 1). For instance, in
Boc-Ala-(S)-Imi-Phe-Val-OMe (2b) the resonance of AlaH appeared at about 5.2 ppm, while
PheH and ValH appeared at about 4.6 and 4.2 ppm.
1
Variable-temperature (VT) H NMR experiments have been widely utilized as a tool for
structure investigation in polypeptides. Usually, for proteins or “rigid” peptides, the H-bonded
amide-NH signals are characterized by modest temperature gradients in absolute value, i.e.
|NH/t| ≤ 2.5 ppb/K,29 while solvent-exposed NHs have larger negative values. Unfortunately, for
acyclic peptides this “rule” has many exceptions.30 Starting from these assumptions, we analyzed
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the NH/t of 0.01 M 2b-e, in 8:2 mixture of [D6]DMSO/H2O (Table S1 and Figure 1). For Boc-
Ala-(S)-Imi-Phe-Val-OMe (2b), the comparatively much lower /t absolute value of Phe3NH (-
0.9 ppb/K) with respect to Ala1NH and Val4NH (-3.7, and -4.5 ppb/K, respectively), indicatively
suggested that the former might be involved in a H-bond. On the other hand, peptides 2c,d showed
high |/t| values for all amide-NHs. Unexpectedly, Ala2NH in Boc-Val-Ala-(R)-Imi-Phe-OMe
(2e) appeared to be significantly less sensitive to increasing temperature than Val1NH and Phe4NH
(/t = -2.4 versus -6.5 and -5.2 ppb/K, respectively), which might be compatible with a
preference for a conformation having AlaNH involved in a H-bond.
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The model compounds were analyzed by 2D-ROESY in 8:2 [D6]DMSO/H2O, using 0.01 M
peptide concentration, and the resulting cross-peaks were ranked by the intensity to infer plausible
interproton distances. Compared to the homochiral Imi-peptides 2b and 2d, the heterochiral
sequences 2c and 2e showed several inter-residue proton-proton correlations between non-
consecutive residues (Figure 1 and Supporting Information).
Ph
Ph
O
O
H
H
O
HN
OMe
(S)
N
OMe
N
H
HN
O
N
H
(S)
N
H
H
O
O
N
H
-0.9
O
H
O
O
N
H
O
H
N
H
2d
2b
H
Boc
N
Boc
Ph
O
O
OMe
Ph
(R)
H
N
HN
OMe
H
N
H
O
H
O
N
(R)
O
H
N
H
HN
O
O
O
H
N
H
N
-2.4
O
O
H
H
H
N
H
2e
N
2c
Boc
Boc
Figure 1. Sketches of 2b-e showing selected, meaningful proton-proton NMR correlations
indicated by arrows. The nonconventional ImiC=O⋯H-C(-1) intramolecular interaction is
rendered in green, strong ROESY correlations are given in blue, medium-intensity correlations in
grey. The /t values for selected amide protons are also shown as red figures.
The estimated distances were analyzed by simulated annealing and restrained molecular
dynamics simulations31 using the AMBER force field32 in explicit water as solvent. In brief, random
geometries of each peptide were obtained by high-temperature unrestrained molecular dynamics
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simulation in a box of standard TIP3P models of equilibrated water.33 For each random structure,
the inter-proton distances deduced by ROESY were introduced as constraints. Neither H-bond
interactions, nor torsion angle restraints were introduced, while  bonds were restricted at 180°,
since the absence of Hi-H(i+1) cross-peaks excluded the occurrence of cis peptide bonds.34 Then
the structures were subjected to high-temperature restrained molecular dynamics with a scaled
force field, followed by a simulation period with full restraints. After slowly cooling the boxes, the
geometries were minimized and the backbones of the Imi-peptides were clustered by the rmsd
analysis.31 In all cases, this procedure gave one major cluster comprising the majority of the
structures (Figure S1). The representative conformers with the lowest energy are reported in Figure
2.
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Figure 2. Representative conformers of 2b-e calculated by ROESY-restrained in a 30×30×30 Å
box of standard TIP3P models of equilibrated water. Heavy dotted lines represent H-bonds, light
dotted grey lines indicate distances; / dihedral angles of selected residues are also reported in
degrees.
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Among the four peptide models, the homochiral structures 2b and 2d showed comparatively
more extended backbone conformations, as expected on the basis of the negligible number of inter-
residue ROESY cross-peaks (Figure 1). Yet, both showed a clear inverse -turn centered on the Imi
residue (Figure 2), and in particular for peptide 2b, having its Imi residue at the position 2 of the
sequence, the turn was stabilized by an explicit H-bond with a H to O distance of 2.0 Å (Figure 2
and Figure 3), consistent with the evidence of VT NMR (Table S1 and Figure 1).
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To simulate the dynamic behavior of the peptides, the ROESY-derived conformers were
analyzed by unrestrained molecular dynamics in explicit water, for 10 ns at rt. During the
simulations, the -turn of 2b resulted to be remarkably stable, correlated to the highly rigid Ala-Imi
central scaffold and the flat imidazolidine-2-one ring. For 2d, having Imi at position 3, the
formation of a definite H-bond was prevented by a certain conformational freedom of the short Phe-
OMe portion, but the -turn appearance was in general maintained. These results were not
unexpected; also peptides containing the homochiral oxazolidin-2-one scaffold showed the
preferential formation of -turns in [D6]DMSO or other competitive solvents.25
As for the heterochiral 2c and 2e, due to the many inter-residue ROESY cross-peaks (Figure
1), the calculated structures were tightly folded (2c and 2eA, Figure 2). The sequence 2c, containing
Imi at the position 2, presented a -turn centered on Imi (Figure 2). Unexpectedly, the ROESY-
derived structure A of 2e (Figure 2) showed a regular -turn centered on Imi, plus an atypical turn
involving the residues Ala-(R)-Imi-Phe, which seemed compatible with a -turn.35
The conformer 2eA was analyzed by unrestrained molecular dynamics, and the analysis of
the trajectories revealed the formation of a 11-membered macrocycle closed by an intramolecular
H-bond between the Ala2NH and Phe4C=O, which is in agreement with the VT NMR temperature
coefficient of AlaNH (Table S1). The representative structure 2eB shown in Figure 2 is
characterized by a distance of 2.4 Å between Ala2C=O and Phe4NH, and of 2.3 Å between Ala2NH
and Phe4C=O (Figure 2 and Figure 3).
The -turn is a rare kind of secondary structure which can be observed in oligo-
cyclopeptides and more seldom in linear peptides and proteins composed of all -amino acids.35
Unlike the more common 5→1 13-membered -, 4→1 10-membered -, and 3→1 7-membered -
turns, the -turn involves 3 residues to form a hydrogen-bonded structure encompassing 11 atoms in
the opposite 2→4 direction (Figure 3A). The / dihedral angles found in 2eB (Figure 2) assign the
-turn to the all-trans class A in the convention of Toniolo and Balaram:35 (/ in degrees) Ala, -
144/+68; (R)-Imi, +91/-47; Phe, -133/+160.
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H
N
Ph
O
H
O
O
H
OMe
O
N
N
(R)
HN
O
N
H
(S)
N
O
N
O
-turn
H
H
Ph
OMe
9
O
'-turn
N
O
H
O
10
11
12
13
-turn
2eB
NH
2b
Boc
HN
Boc
-turn
A
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O
R1
H
O
R4
4
N
H
N
2
N
1
3
N
R
H
O
R2
H
O
-turn
-turn
C
D
B
O
N
O
-turn
H
(R)
N
H
O
O
H
N
O
O
H
N
(R)
O
OtBu
N
-turn
H
O
O
Figure 3. Sketch of the expected inverse -turn of the homochiral 2b; the ImiC=O⋯H-C(-1)
interaction is rendered in green. Sketch of the class A /-mixed turns35 in 2eB. A) “Common way”
intramolecular hydrogen bonds in a tetrapeptide (residues numbered 1 to 4) rendered as continuous
grey arrows, and atypical 11 membered -turn in the 2→4 reversed direction as a dotted arrow. B)
Oligomers composed of Ala-oxazolidin-2-one units produce a helix of “normal” -turns.3c-25 C)
Crystal structure of the oxazolidin-2-one25c and D) of the imidazolidin-2-one scaffolds.23
The comparison between the structures of 2c and 2e in Figure 2 showed that the -turn was
not completely formed in 2c, albeit both compounds share the same Ala-(R)-Imi-Phe tripeptide
sequence. Possibly, the failure of 2c in forming a unequivocal H-bonded 2→4 turn can be attributed
to presence of the bulky Boc group at Ala1, as suggested by the analysis of the trajectories of the
unrestrained molecular dynamics, which showed the clash between the tBu and Val4.
Finally, it is worth mentioning that in spite of the structural similarity with (R)-Imi, the (R)-
oxazolidin-2-one-4-carboxylate scaffolds produced only “normal” conformations, i.e. -turns in
[D6]DMSO and -turns in CDCl3.25a-b And oligomers formed of repetitive Ala-(R)-oxazolidin-2-
one-4-carboxylate dipeptide units (≥ 5) folded in a ordered structure to give a variant of the 310-
helix, namely a series of consecutive regular 4→1 -turns (Figure 3B), in the crystal state and in
MeOH.25c At present, the exhaustive investigation of the structural features of the Imi scaffold is
beyond the scope of this work. Nevertheless, some clues can be perceived by the inspection of the
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X-ray crystal structures of 3-acyl-oxazolidin-2-one-4-carboxylates25c and 3-acyl-imidazolidine-2-
one-4-carboxylates23 (Figure 3C,D). The comparison highlighted some relevant 3D differences
which might be responsible for the different H-bonding patterns. While the geometry of the 5-
membered rings is quite twisted in the oxazolidin-2-one (Figure 3C),25c it is extraordinarily flat in
the Imi ring (Figure 3D).23 This necessarily translates into different orientations of C4-carboxylate
vectors, reasonably fundamental to the development of the overall conformations, and in particular
the carboxylate group in the oxazolidin-2-one adopts an axial, almost vertical orientation above the
5-membered ring (Figure 3C).
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Imidazolidin-2-ones substituted at the position 1. The formation of the Imi ring was attained by
sacrificing the N-amino group of Dap. Hence, Imi can be regarded to as a pseudo Pro showing the
sole trans conformation at the preceding peptide bond. Intriguingly, conformational analysis by
NMR and molecular dynamics has shown that (S)-Imi peptides adopt an inverse -turn
conformation, while (R)-Imi promotes the formation of the rare - secondary structure. This
unexpected observation prompted us to possibly expand the scope of the Imi rings, making possible
their use as general promoters of such unusual secondary structures also in sequences having at the
position i residues other than a (pseudo) Pro. For this reason, we tackled the opportunity to
introduce any substituents at the position 1 of the Imi scaffolds.
The strategy we adopted is outlined in Scheme 3. Dipeptides containing (S)- or (R)-serine 3
were prepared in solution as described in the literature.36 The dipeptide esters 3 were isolated in
good yield (in the 70-85 % range, 80-90 % pure according to analytical RP HPLC) by flash
chromatography over silica gel. The dipeptides 3 were treated with a mixture of triphenylphosphine,
iodine (1.2 equiv each), and imidazole (2.5 equiv),37 affording -iodo-Ala-dipeptides 4, not isolated
due to intrinsic instability;37 TLC analyses of the reaction mixtures revealed the complete
consumption of 3.
The displacement of iodine with alkyl or arylamines, i.e. methylamine, isopropylamine,
aniline, allylamine, -aminobutyric acid (GABA) methyl ester (2.0 equiv.) in DMF gave the N-
substituted Dap dipeptides 5. Again, the TLC analyses of the crude reaction mixtures showed the
disappearance of the reagents. Unfortunately, with the exception of Boc-Ala-Dap(N-allyl)-OMe 5d,
the isolation of these dipeptides 5 was not possible, either by chromatography over silica gel or by
preparative RP HPLC, for the accumulation of the many side-products.
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H
N
I
R3
OH
OR2
O
PPh3 (1.2 eq),
I2 (1.2 eq)
O
O
H
R3-NH2
H
H
carbonate
base
N
OR2
N
OR2
N
R
N
R
N
R
N
H
H
R1
(2.0 eq)
H
R1
imidazole
(2.5 eq)
O
R1
O
O
7
8
3
4
5
R4
O
9
R4 = succinimidyl-O
Cl
R3
O
O
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11
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N
R1
R3
N
O
O
tBuO
R
N
H
N
N
OR2
MeO
OR2
R
N
H
imidazolyl-O
p-nitropheny-O
H
R1
O
7
6
(S)-Imi, R = Boc, R1 = R2 = R3 = Me
(R)-Imi, R = Boc, R1 = R2 = R3 = Me
a
b
c
d
e
f
(S)-Imi, R = Cbz, R1 = R3 = iPr, R2 = Me
(S)-Imi, R = Boc, R1 = Me, R2 = Bn, R3 = Ph
(S)-Imi, R = Boc, R1 = Me, R2 = Me, R3 = allyl
(S)-Imi, R = Boc, R1 = Me, R2 = Bn, R3 = allyl
(R)-Imi, R = Cbz, R1 = R3 = iPr, R2 = Me
g
h
i
(R)-Imi, R = Boc, R1 = Me, R2 = Bn, R3 = Ph
(R)-Imi, R = Boc, R1 = Me, R2 = Bn, R3 = allyl
(S)-Imi, R = Boc, R1 = Me, R2 = Bn, R3 = (CH2)3CO2Me
l
Scheme 3. Synthesis of N1-substituted Imi-peptides 7.
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
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40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Hence, we opted for the direct cyclization of the crude dipeptides 5 with DSC/DIPEA, as
reported above for the synthesis of Imi-peptides 2. This reaction gave the formation of the
intermediate N-carbamate 6, and of desired 7 only in traces, as observed by RP HPLC and ESI MS
analysis of the reaction mixture. Even the highly pure 5d failed to give the corresponding 7d,
suggesting that the scarce purity of the other reagents 5 was not the only responsible for the
disappointing result. Thereafter, the procedure was repeated upon varying the conditions, using as
model reagent the crude dipeptide Boc-Ala-Dap(N1Me)-OMe (5a), obtained from the Boc-Ala--
iodoAla-OMe 4a and methylamine. Besides to DIPEA, also Na2CO3, DMAP, and DBU, were
tested. The increase of the amounts of DSC and/or of the base, the increase of temperature, and of
reaction time, had very little impact (not shown), while the use of DMF as the only solvent gave a
modest increase of the yield (Table 2, entry 1).
Apart from DSC, the use of other carbonates or dicarbonates such as triphosgene, Boc2O,
methyl chloroformate, CDI, entries 2 to 5, respectively, in combination with DIPEA or the other
bases as above, was also poorly effective. Gratifyingly, p-nitrophenylchloroformate (pNPC)38 and
DIPEA (1.5 equiv each) in DMF at rt after 12 h gave 7a in 70 % yield over 3 steps, accompanied by
the N-p-nitrophenoxycarbonyl-Dap(Me) intermediate 6 (25 %) (entry 6). The yield was further
increased by changing DIPEA with DBU, and 7a was isolated in excellent amount (90 %) by flash
chromatography over silica gel (entry 7).
The same conditions were applied to the crude dipeptides 5b,c,e, obtained as reported in
Scheme 3 from the corresponding -iodo-derivatives 4 and isopropylamine, aniline, allylamine,
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respectively. The N-terminal capping group, the ester, and the amino acid preceding Dap, were
varied to check the generality of the process. The benzyl ester 5e was preferred to the methyl ester
5d for the convenience of further transformations, as discussed in the next sections. In all cases, the
reactions gave results comparable to 5a (entries 8 to 10).
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Table 2. Formation of imi-dipeptides 7 using carbonate/base (1.5 equiv) in DMF, after 12 h at rt.
entry
1
compd
Carbonate/base
7 (%)
<10b
traces
“
purity (%)a
-
6 (%)
a
“
“
“
“
“
“
b
c
e
f
DSC/DIPEA
70b
d
d
2
(Cl3CO)CO/basec
-
-
3
Boc2O/basec
-
75
4
ClCO2Me/basec
“
-
80
5
CDI/basec
<5b
70b
90e
85e
80e
93e
87e
82e
77e
90e
74e
-
Traces
6
pNPC/DIPEA
-
25
7
pNPC/DBU
95
91
92
94
93
91
96
92
94
traces
8
“
“
“
“
“
“
“
“
“
9
10b
10
11
12
13
14
15
traces
“
g
h
i
“
11b
“
l
“
a
b
After isolation, determined by RP HPLC ESI MS. Determined by the RP HPLC analyses of the reaction
c
d
e
mixtures. Base: DIPEA, carbonate, DMAP, DBU. Complex mixture of by-products. Determined after
isolation by flash chromatography over 3 synthetic steps respect to peptides 3.
Subsequently, the procedure was repeated starting from the dipeptides containing D-serine.
As for the (S)-configured series, the formation of the dipeptides containing (R)--iodoAla, followed
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by displacement with the same amines, gave the dipeptides containing N-methyl, isopropyl, phenyl,
and allyl (R)-Dap. Then, the treatment with pNPC/DBU gave (R)-Imi peptides 7f, 7g, 7h, and 7i in
good to excellent yields (entries 11 to 14).
8
9
Finally, the dipeptide Boc-Ala-(S)-Imi[(CH2)3CO2Me]OBn (7l) (Scheme 3), was prepared in
sufficient yield by treatment of the -iodoAla-dipeptide with GABA methyl ester, followed by
cyclization under the usual conditions (entry 15).
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Epimerization during the displacement of iodine from -iodoAla by the amines, or during
1
the following cyclization step, was excluded based on the comparison of the H NMR of the
(S,S)/(S,R) diastereomeric pairs with the peptides 2b-e containing unsubstituted Imi. In particular,
for all compounds containing (R)-Imi in CDCl3 the proton ImiH4 appeared ≤ 4.75 ppm, while for
compounds containing (S)-Imi ImiH4 appeared ≥ 4.8 ppm.
In any case, to confirm that the stereochemistry was maintained, the diastereomeric 7a and
7f were treated with LiOOH in THF/water at rt for 4 h, followed by 1M HCl,39 leading to the
detachment of Boc-Ala-OH and the release of the enantiomeric methyl 1-methyl-2-
oxoimidazolidine-4-carboxylates (S)-8 and (R)-8. After purification by flash chromatography over
silica gel (eluent cyclohexane/EtOAc 1:3) their specific optical rotation was determined (Scheme 4)
and judged in agreement with the literature for (S)-8: []D20 +26.9 (c 1, MeOH).23
8
(S)-
80%
N
[]D20 + 25
O
O
N
O
LiOOH
THF/H2O
(c 0.7, MeOH)
N
HN
(S)
O
Boc
N
O
O
H
O
8
(R)-
75%
[]D20 - 24
a
f
7
(S)-Imi
(R)-Imi
(c 0.6, MeOH)
Scheme 4. Cleavage of the Imi rings 8 from 7a,f.
These Imi dipeptide scaffolds carrying three possible different pharmacophores can be of
some interest in peptidomimetic chemistry. There is evidence for the optimality of three-residue
motifs for biological activity.40 In a similar way as shown for trans-pyrrolidine-3,4-dicarboxamide
templates by D. L. Boger et al. (Figure 4A),41 the triply functionalized Imi dipeptides 7 (Figure 4)
might be regarded as potential -turn mimetics when the turn amino acid i+1 serves as a structural
rather than a recognition role, e.g. Pro or Gly (Figure 4B). Hence, Imi scaffolds reproducing the
side chains of the residues i, i+2, i+3, could still maintain the bioactivity of the native peptides.
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As for the oligomers, the Imi scaffolds have been shown to act as proline mimics inducing
all-trans conformations in the sequences, and to promote classic or unusual turn structures,
depending on the stereochemistry. Interestingly, while most of the conformationally restricted turn-
inducing elements (e.g. pseudoprolines, Freidinger lactams, spirolactams, bicyclic thiazolidines,
etc.) are generally deprived of some relevant pharmacophores,1,42 the Imi rings can be equipped
with proteogenic side chains (Figure 4C), e.g. of Ala, Val, Phe (Scheme 3).
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R2
R2
O
O
O
O
H
N
O
R3
N
O
N
N
R3
HN
R
R
O
A
R2
7
i+2
O
O
i+1
HN
N
H

i+3
N

R3
H
O
R
i
B
R1
N
O
R2
H
CO2Me
N
N
N
O
H
N
O
R3
O
C
N
O
Boc
OBn
N
R
H
H
O
O
N
7l
Figure 4. A) Pyrrolidine- and Imi-based -turn mimetics 7.41 B) Classic - or -turns. C) Imi as a
turn inducing element for peptidomimetics equipped with side chains at each residue. Triply
substituted Imi peptide 7l with orthogonally-protected, further functionalizable groups.
To further challenge the potential utility of the N1-substituted Imi dipeptides, Boc-Ala-(R)-
Imi(N1allyl)-OBn (7i) was subjected to catalytic hydrogenation, and the resulting (R)-
.
Imi(N1propyl) dipeptide acid was coupled with H-Phe-OMe HCl, giving the tripeptide 9 equipped
1
with a non proteogenic side chain (Scheme 5). As expected, the H NMR analysis of 9 in 8:2
[D6]DMSO/H2O supported that this Imi-N1-propyl tripeptide adopted a conformation consistent to
that of the heterochiral models 2c or 2e. Indeed, the 1H NMR of 9 was practically superimposable to
that of the Boc-Ala-(R)-Imi-Phe portion of 2c, while the only difference with the corresponding
portion in 2d was the obviously different chemical shift of AlaNH (Supporting Information). See
for instance the highly reproducible pattern of the resonances of H4 and the two H5 protons
(Supporting Information).
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N
O
1. H2 Pd/C
N
Boc
(R)
OBn
2. HOBt (1.2 eq)
EDC (1.2 eq)
TEA (2.5 eq)
N
H
O
O
O
.
H-Phe-OMe HCl (1.2 eq)
7i
8
9
10
11
12
13
14
15
16
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19
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22
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24
25
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58
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60
N
9
78%
H
N
Boc
(R)
N
N
CO2Me
H
O
O
Ph
Scheme 5. Synthesis of the N1-substituted Imi-tripeptide 9 from the N1-allyl Imi-tripeptide 7i.
On the other hand, the synthetic strategy described above was utilized also for introducing
substituents carrying orthogonally protected functional groups. A first example was the dipeptide
Boc-Ala-(S)-Imi[(CH2)3CO2Me]OBn (7l) (Table 2, entry 15, and Figure 4), prepared with GABA
methyl ester. In perspective, after hydrolysis under basic conditions, the N1-butanoic acid side chain
might be exploited to graft the peptide onto e.g. surfaces, polymeric materials, nanoparticles, etc.
Finally, the N1-substituent of Imi can be equipped with a tag-residue for site-selective
modifications of the sequences. Among the most popular reactions utilized to transform such
handles without perturbing the remaining amino acids, it is possible to cite the Cu-catalyzed azide-
alkyne cycloaddition, the Sonogashira, Suzuki-Miyaura, Mizoroki-Heck, Diels-Alder reactions, the
photo-1,3-dipolar cycloaddition, the Staudinger ligation, and the olefin metathesis with Grubbs or
Hoveyda-Grubbs catalysts.17
As a proof of concept, the allyl substituent at the position 1 of the Imi ring in 7e was
selectively derivatized with N-Boc-4-bromoaniline by the Heck reaction (Scheme 6). Conducted in
DMF in the presence of Pd(OAc)2/PPh3/TEA, under inert atmosphere, at 80°C,43 the reaction gave
the adduct 10 characterized by a exclusively trans connection between the two portions, as
determined by the -CH=CH- coupling constants in the 1H NMR, J = 15.6 Hz (for trans alkenes J =
12-18 Hz, for cis alkenes J = 6-12 Hz). The reduction by catalytic hydrogenation gave a fully
flexible linkage, and at the same time removed the benzyl ester protection, giving the dipeptide acid
11, ready in prospective for the extension of the peptidic backbone.
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H
N
H
N
Boc
Boc
5
6
7
8
N
H
N
O
Pd(OAc)2 (0.01 eq),
PPh3 (0.02 eq),
Boc
H2 Pd/C
N
10
78%
11
99%
N
N
OBn
+
O
O
Br
TEA/DMF,
N2, 80 °C
O
Boc NH
O
N
N
1.5 eq
9
OH
OBn
Boc
Boc
N
N
7e
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
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51
52
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O
O
O
O
H
H
Scheme 6. Heck reaction of the N1-allyl Imi-tripeptide 7e, giving 10 and then 11.
Conclusions
The introduction of nitrogen heterocycles into peptide sequences represents an effective approach to
increase enzymatic stability and to induce well-defined secondary structures. To this purpose, we
developed an expedient synthesis of hybrid peptides containing imidazolidin-2-one-4-carboxylate
(Imi) scaffolds. These exceptionally flat pseudo-prolines demonstrated the ability to favor unusual
conformations in a competitive and biomimetic solvent. While (S)-Imi peptides adopted a -turn
conformation, (R)-configured Imi promoted the additional formation of a -turn, an infrequent
secondary structure characterized by a 11-membered 2→4 H-bond going opposite respect to the
classic - or -turns. Besides, Imi can be equipped at the position N1 with substituents that
reproduce the side chains of native amino acids, including Ala, Val, or Phe, in either L- or D-
configuration, as well as with non-proteinogenic substituents, such as n-propyl or allyl. In this
respect, the N1-substituted (R)-Imi rings represent a new class of general promoters the rare -turn
in a variety of peptide sequences. On the other hand, Imi can also carry functional groups, for
instance the allyl group, capable of selective reactions without perturbing the rest of the structure,
e.g. by the Heck reaction. Potential future applications might include the glycosylation, prenylation,
PEGylation, biotinylation, or attachment to solid surfaces, self-assembled monolayers, proteins, or
the conjugation with fluorophores or antibodies.17,44
Experimental Section
General Experimental Methods. All purchased reagents were used without further purifications.
Purities were assessed by analytical RP HPLC, and confirmed by exact mass, and elemental
analysis. MW lab station: MicroSYNTH equipped with a built-in ATC-FO advanced fiber optic
automatic temperature control. RP HPLC: Agilent 1100 series apparatus, with a RP column
Phenomenex mod. Gemini 3 C18 110 Å 100×3.0 mm, stationary phase octadecyl carbon chain-
bonded silica with TMS endcapping, fully porous organo-silica solid support, particle size 3 m,
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pore size 110 Å, length 100 mm, internal diameter 3 mm, DAD 210 and 254 nm, mobile phase from
9:1 to 2:8 water/CH3CN, in 20 min, at a flow rate of 0.5 mL min-1, followed by 10 min at the same
composition. Semi-preparative RP-HPLC: Agilent 1100 series, RP column ZORBAX mod. Eclipse
XDBC18 PrepHT cartridge 21.2×150 mm 7, stationary phase octadecyl carbon chain bonded
silica, double endcapped, particle size 7 mm, pore size 80 Å, length 150 mm, internal diameter 21.2
mm, DAD 210 nm; mobile phase from H2O/CH3CN (8:2) to CH3CN (100 %) in 10 min at a flow
rate of 12 mL min-1. HRMS: Waters Xevo QTOF. Routine ESI MS: MS single quadrupole HP
1100MSD detector, drying gas flow of 12.5 L min-1, nebulizer pressure 30 psgi, drying gas temp
350°C, capillary voltage 4500(1) and 4000(2), scan 50-2600 amu. Elemental analysis: Thermo
Flash 2000 CHNS/O. NMR characterization was done on a Varian Gemini 400 or 200, using 0.01-
0.04 M peptide in 5 mm tubes at rt, using the solvents CDCl3, or [D6]DMSO, or 8:2
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1
[D6]DMSO/H2O (water suppression by the solvent presaturation procedure PRESAT); H spectra
were recorded at 400 MHz, and the unambiguous assignment of the resonances was done by 2D
gCOSY; proton-decoupled 13C NMR were recorded at 100 or 75 MHz; chemical shifts were
reported as  values in ppm relative to solvent as internal standard.
Synthesis of peptides 1 in solution. General Procedure. A mixture of the Boc-amino acid (0.3
mmol) and HOBt (0.049 g, 0.36 mmol) was stirred in 3:1 DCM/DMF (5 mL) at rt, and after 10 min
.
the amino ester counterpart (0.36 mmol), EDCI HCl (0.069 g, 0.36 mmol) and TEA (0.13 mL, 0.94
mmol) were added in sequence while stirring at rt. After 2 h, the solvents were removed at reduced
pressure, and the residue was diluted with EtOAc (20 mL). The slurry mixture was washed with 0.1
M HCl (5 mL), and a saturated solution of NaHCO3 (5 mL). The organic layer was dried over
Na2SO4 and the solvent was evaporated at reduced pressure. The intermediate crude peptides,
obtained in quantitative yield, were analyzed by RP HPLC and ESI MS, and were used without
further purifications.
Boc deprotection was accomplished by stirring the crude peptides in 1:3 TFA/DCM (4 mL)
for 30 min. Then the mixture was concentrated at reduced pressure, and the treatment was repeated.
The residue was suspended in Et2O (20 mL), and the peptide-TFA salts which precipitated in almost
quantitative yield were used for the next couplings without further purifications.
The final sequences 1 were isolated by flash chromatography over silica-gel (eluent 1:2
cyclohexane/EtOAc), 80-85 % yield, 80-90 % pure by RP HPLC (General Methods). The identity
was confirmed by ESI MS analysis (General Methods).
Cyclization to Imi-peptides 2a-c in solution. General Procedure. The peptides 1 (0.2 mmol) were
dissolved in EtOH (15 mL), and a 2 L balloon filled with H2 was applied in the presence of a
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catalytic amount of 10 % Pd/C, while stirring at rt. The reactions were monitored by TLC, and were
generally judged complete in 6 h. Then the mixture was filtered over celite® and the solvent was
evaporated at reduce pressure.
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The residue was dissolved in in 3:1 DCM/DMF (5 mL) and DSC (0.061 g, 0.24 mmol) and
DIPEA (0.044 mL, 0.24 mmol) were added at rt under inert atmosphere. The mixture was stirred for
3 h, then the solvent was distilled under reduced pressure, the residue was diluted with 0.1 M HCl
(5 mL), and the mixture was extracted three times with DCM (10 mL). The combined organic
layers were dried over Na2SO4, and concentrated at reduced pressure. The residue was separated by
semi-preparative RP HPLC on a C18 column, eluent from H2O/CH3CN (8:2) to CH3CN (100 %) in
10 min, flow rate 12 mL min-1 (General Methods), and purity was determined by RP HPLC
(General Methods).
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Boc-Ala-(S)-Imi-Phe-OMe (2a). Cyclization of 1a (0.11 g, 0.19 mmol) according to the General
1
Procedure gave 2a (0.077 g, 86 %, 92 % pure). H NMR (8:2 [D6]DMSO/H2O, 400 MHz)  1.17
(d, J = 7.1 Hz, 3H, AlaMe), 1.35 (s, 9H, tBu), 2.93-3.02 (m, 2H, PheH), 3.08 (dd, J = 3.0, 9.6 Hz,
1H, ImiH5), 3.57 (s, 3H, OMe), 3.63 (dd, J = 9.6, 10.0 Hz, 1H, ImiH5), 4.44 (m, 1H, PheH), 4.75
(dd, J = 3.0, 10.0 Hz, 1H, ImiH4), 5.18 (dd, J = 7.1, 14.4 Hz, 1H, AlaH), 7.09 (d, J = 8.0 Hz, 1H,
AlaNH), 7.18-7.35 (m, 5H, ArH), 7.73 (s, 1H, ImiNH1), 8.58 (d, J = 6.8 Hz, 1H, PheNH); 13C{1H}
NMR ([D6]DMSO, 100 MHz),  18.2, 29.1, 37.5, 41.3, 49.0, 52.8, 54.7, 55.3, 78.8, 127.6, 129.3,
130.0, 137.7, 156.1, 170.6, 172.7, 174.4; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for
C22H31N4O7 463.2193; Found 463.2164. Anal. Calcd for C22H30N4O7: C, 57.13; H, 6.54; N,
12.11. Found: C, 57.19; H, 6.49; N, 11.99.
Boc-Ala-(S)-Imi-Phe-Val-OMe (2b). Cyclization of 1b (0.14 g, 0.21 mmol) according to the
General Procedure gave 2b (0.10 g, 88 %, 93 % pure). 1H NMR (8:2 [D6]DMSO/H2O, 400 MHz) 
0.82-0.93 (m, 6H, ValMe), 1.17 (d, J = 7.2 Hz, 3H, AlaMe), 1.34 (s, 9H, tBu), 2.01 (m, 1H,
ValH), 2.80 (dd, J = 8.8, 13.4 Hz, 1H, PheH), 2.99 (dd, J = 4.0, 13.4 Hz, 1H, PheH), 3.08 (dd, J
= 2.2, 9.8 Hz, 1H, ImiH5), 3.58 (m, 1H, ImiH5), 3.62 (s, 3H, OMe), 4.14 (dd, J = 7.2, 7.6 Hz, 1H,
ValH), 4.56 (m, 1H, PheH), 4.73 (dd, J = 2.8, 7,2 Hz, 1H, ImiH4), 5.16 (m, 1H, AlaH), 7.00 (d,
J = 7.6 Hz, 1H, AlaNH), 7.16-7.30 (m, 5H, PheArH), 7.71 (s, 1H, ImiNH1), 8.24 (d, J = 7.6 Hz, 1H,
ValNH), 8.28 (d, J = 8.0 Hz, 1H, PheNH); 13C{1H} NMR (CDCl3, 100 MHz)  17.8, 18.8, 28.3,
29.7, 31.1, 38.1, 49.0, 52.2, 54.9, 56.0, 57.5, 79.9, 127.1, 127.2, 128.8, 129.1, 129.3, 136.0, 154.6,
155.2, 168.5, 170.2, 171.6, 175.0; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for C27H40N5O8
562.2877; Found 562.2821. Anal. Calcd for C27H39N5O8: C, 57.74; H, 7.00; N, 12.47. Found: C,
57.81; H, 6.90; N, 12.54.
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Boc-Ala-(R)-Imi-Phe-Val-OMe (2c). Cyclization of 1c (0.12 g, 0.18 mmol) according to the
General Procedure gave 2c (0.092 g, 91 %, 93 % pure). 1H NMR (8:2 [D6]DMSO/H2O, 400 MHz)
 0.88 (d, J = 7.2 Hz, 3H, ValMe), 0.91 (d, J = 6.8 Hz, 3H, ValMe), 1.15 (d, J = 6.4 Hz, 3H,
AlaMe), 1.35 (s, 9H, tBu), 2.05 (m, 1H, ValH), 2.42 (m, 1H, ImiH5), 2.71 (dd, J = 10.0, 13.7 Hz,
1H, PheH), 3.06 (dd, J = 3.8, 13.7 Hz, 1H, PheH), 3.34 (dd, J = 4.8, 12.0 Hz, 1H, ImiH5), 3.63
(s, 3H, OMe), 4.30 (m, 1H, ValH), 4.56 (m, 1H, ImiH4), 4.71 (ddd, J = 3.8, 8.8, 10.0 Hz, 1H,
PheH), 5.31 (dq, J = 6.4, 8.8 Hz, 1H, AlaH), 6.62 (d, J = 8.8 Hz, 1H, AlaNH), 7.17-7.27 (m, 5H,
PheArH), 7.62 (s, 1H, ImiNH1), 8.20 (d, J = 7.6 Hz, 1H, ValNH), 8.37 (d, J = 8.8 Hz, 1H, PheNH);
13C{1H} NMR ([D6]DMSO, 100 MHz)  18.5, 19.0, 19.2, 28.3, 29.9, 30.8, 38.0, 48.1, 51.8, 53.0,
55.2, 57.8, 78.1, 126.4, 128.0, 129.4, 137.6, 154.6, 155.0, 169.2, 171.3, 171.8, 172.9; HRMS
(ESI/QTOF) m/z: [M + H]+ Calcd for C27H40N5O8 562.2877; Found 562.2849. Anal. Calcd for
C27H39N5O8: C, 57.74; H, 7.00; N, 12.47. Found: C, 57.55; H, 7.13; N, 12.17.
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Synthesis of Imi-peptides 2d,e in solid-phase. General Procedure. Wang resin pre-loaded with
Fmoc-Phe (0.25 g, 0.4-0.8 mmol g−1, particle size 100-200 mesh) was placed into a reactor
equipped with a filter, and suspended in DCM (5 mL). Fmoc was removed by treatment with 1:4
piperidine/DMF (5 mL) under MW irradiation at 40 W while bubbling N2 in an open vessel,
monitoring the internal temperature at 45°C. After 2 min, the suspension was filtered, and the
procedure was repeated. Then, the resin was washed three times in sequence with DCM, DMF, and
MeOH (5 mL each).
The resin was swollen in DCM (8 mL), and a mixture of Fmoc-Dap(Bn)OH (0.137 g, 0.3
mmol) and HOBt (0.041 g, 0.3 mmol) in DMF (4 mL) was added, followed by DCC (0.078 g, 0.38
mmol). The mixture was heated at 45°C for 10 min under MW irradiation as described above, while
bubbling N2. The resin was filtered and washed three times with the sequence DCM, DMF, MeOH
(5 mL each). Coupling efficacy was determined by means of the Kaiser test. All the remaining
residues were attached by the same protocols.
The resulting peptidyl-resin 1d or 1e was suspended in 1:3 iPrOH/toluene (10 mL), and
HCO2NH4 (0.038 g, 0.6 mmol) and a catalytic amount of 10 % Pd/C were added in sequence. The
mixture was irradiated at 600 W for eight cycles of 1 min each. The resin-bound peptide was
filtered and washed three times in sequence with DCM, DMF, and MeOH (5 mL each).
Then the peptidyl-resin was suspended in 3:1 DCM/DMF (5 mL), and DSC (0.116 g, 0.45
mmol) and DIPEA (80 L, 0.45 mmol) were added at rt under inert atmosphere. After 3 h the
mixture was filtered, and the resin-bound peptide was washed three times in sequence with DCM,
DMF, and MeOH (5 mL each).
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A suspension of the peptidyl-resin in 1:3 triethylamine/MeOH (20 mL) was heated at 60°C
for 20 min under MW irradiation in an open vessel. After filtration, the resin was washed twice with
DCM and Et2O (10 mL each), the filtrates were collected, and the volatiles were evaporated at
reduced pressure. The Imi-peptides were separated by semi-preparative RP HPLC on a C18 column
(General Methods) giving 2d or 2e in good yield (calculated on an average resin load of 0.6 mmol
g−1), and high purity, as determined by analytical RP HPLC (General Methods).
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Boc-Val-Ala-(S)-Imi-Phe-OMe (2d). The procedure gave peptide 2d (0.069 g, 84 %, 95 % pure).
1H NMR (8:2 [D6]DMSO/H2O, 400 MHz)  0.85 (d, J = 6.8 Hz, 3H, ValMe), 0.87 (d, J = 6.8 Hz,
3H, ValMe), 1.24 (d, J = 7.0 Hz, 3H, AlaMe), 1.38 (s, 9H, tBu), 1.93 (m, 1H, ValH), 2.97 (dd, J =
7.6, 13.8 Hz, 1H, PheH), 3.02 (dd, J = 5.6, 13.8 Hz, 1H, PheH), 3.09 (dd, J = 3.4, 9.4 Hz, 1H,
ImiH5), 3.57 (s, 3H, OMe), 3.62 (m, 1H, ImiH5), 3.79 (m, 1H, ValH), 4.44 (ddd, J = 5.6, 6.8, 7.6
Hz, 1H, PheH), 4.75 (dd, J = 3.4, 10.0 Hz, 1H, ImiH4), 5.45 (m, 1H, AlaH), 6.59 (d, J = 9.2 Hz,
1H, ValNH), 7.19-7.24 (m, 3H, PheArH), 7.25-7.31 (m, 2H, PheArH), 7.77 (s, 1H, ImiNH1), 8.00
13
(d, J = 7.6 Hz, 1H, AlaNH), 8.59 (d, J = 6.8 Hz, 1H, PheNH); C{1H} NMR ([D6]DMSO, 75
MHz)  18.5, 19.1, 20.1, 29.1, 29.9, 30.4, 31.5, 37.4, 52.8, 54.7, 55.3, 60.1, 79.9, 127.6, 129.2,
130.0, 137.7, 156.0, 156.7,170.5, 171.7, 172.6, 174.4; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for
C27H40N5O8 562.2877; Found 562.2822. Anal. Calcd for C27H39N5O8: C, 57.74; H, 7.00; N,
12.47. Found: C, 57.85; H, 6.85; N, 12.58.
Boc-Val-Ala-(R)-Imi-Phe-OMe (2e). The procedure gave peptide 2e (0.071 g, 86 %, 96 % pure).
1H NMR (8:2 [D6]DMSO/H2O, 400 MHz)  0.76 (d, J = 6.8 Hz, 3H, ValMe), 0.81 (d, J = 6.0 Hz,
3H, ValMe), 1.20 (d, J = 6.4 Hz, 3H, AlaMe), 1.35 (s, 9H, tBu), 1.94 (m, 1H, ValH), 2.67 (dd, J =
2.2, 9.8 Hz, 1H, ImiH5), 2.88 (dd, J = 10.0, 13.3 Hz, 1H, PheH), 3.06 (dd, J = 5.4, 13.3 Hz, 1H,
PheH), 3.45 (m, 1H, ImiH5), 3.61 (s, 3H, OMe), 3.80 (dd, J = 6.6, 8.0 Hz, 1H, ValH), 4.48 (ddd,
J = 5.4, 8.0, 10.0 Hz, 1H, PheH), 4.60 (dd, J = 2.2, 9.8 Hz, 1H, ImiH4), 5.58 (m, 1H, AlaH), 6.75
(d, J = 8.0 Hz, 1H, ValNH), 7.16-7.22 (m, 3H, PheArH), 7.23-7.30 (m, 2H, PheArH), 7.73 (s, 1H,
13
ImiNH1), 7.86 (d, J = 8.0 Hz, 1H, AlaNH), 8.54 (d, J = 8.0 Hz, 1H, PheNH); C{1H} NMR
([D6]DMSO, 100 MHz)  18.9, 19.2, 19.6, 28.6, 30.7, 37.4, 41.0, 47.1, 52.3, 53.7, 55.4, 56.5, 60.0,
78.6, 124.8, 126.0, 127.0, 128.6, 129.6, 137.4, 155.2, 155.9, 169.6, 171.0, 172.0, 172.7; HRMS
(ESI/QTOF) m/z: [M + H]+ Calcd for C27H40N5O8 562.2877; Found 562.2845. Anal. Calcd for
C27H39N5O8: C, 57.74; H, 7.00; N, 12.47. Found: C, 57.90, H, 6.83, N, 12.60.
Conformational analysis by NMR. Peptide samples were dissolved in 8:2 [D6]DMSO/H2O in 5
mm tubes to the final concentration of 0.01 M. At this concentration, the intramolecular aggregation
in mixtures of [D6]DMSO and H2O is usually unimportant. Besides, self-association of the peptides
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was excluded based on the reproducibility of the chemical shift of non-exchangeable protons in the
concentration range 0.01-0.04 M (not shown). Water suppression was achieved by the PRESAT
procedure implemented in Varian. Proton resonance assignment was accomplished through
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gCOSY. VT H NMR experiments were recorded over the range of 298-348 K; temperature
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calibration was done with the ethylene glycol OHCHn chemical-shift separation method. 2D
ROESY experiments were done at rt, phase-sensitive mode, spin-locking field (b2) = 2000 Hz,
mixing time = 250 ms; spectra were processed in the hypercomplex approach; peaks were
calibrated on solvent. Only ROESY-derived constraints were included in the restrained MD. Cross-
peak intensities were ranked and associated to the distances (Å): very strong = 2.3, strong = 2.6,
medium = 3.0, weak = 5.0. The intensities of the cross peaks arising from protons separated by
known distances (e.g., geminal) were found to match with these associations, but were discarded.
For the absence of H(i, i+1) ROESY cross peaks, all of the  bonds were set at 180° (f constant:
16 kcal mol-1 Å-2).
MD simulations. The restrained MD simulations were conducted at 300 K and 1 atm by using the
AMBER force field in a 30×30×30 Å3 box of standard TIP3P models of equilibrated water, periodic
boundary conditions dielectric scale factor = 1, cutoff for the nonbonded interactions = 12 Å; all
water molecules closer than 2.3 Å to a solute atom were eliminated. 50 Random structures were
generated by a 100 ps simulation at 1200 K; these were subsequently subjected to restrained MD,
50 ps with a 50 % scaled force field at 1200 K, then by 50 ps with full distance restraints, force
constant = 7 kcal mol-1Å-2, after which the system was cooled in 20 ps to 50 K. H-bond interactions
were not included, nor were torsion angle restraints. The resulting structures were minimized by
3000 cycles of steepest descent and 3000 cycles of conjugated gradient, convergence = 0.01 kcal Å-
1mol-1. The backbones of the structures were clustered by the rmsd analysis.
Unrestrained MD simulations were performed starting with the conformation derived from
ROESY in the box of standard TIP3P water for 10 ns at 298 K using periodic boundary conditions,
at constant temperature and pressure (Berendsen scheme,45 bath relaxation constant of 0.2). For 1-4
scale factors, van de Waals and electrostatic interactions are scaled in AMBER to half their nominal
value. The integration time step was set to 0.1 fs. The system coordinates were collected every
picosecond.
Synthesis of peptides 3. The reaction was performed under conditions which allow using (S)- or
(R)-serine esters without protection of the hydroxy group.36 Briefly, a stirred solution of the N-
protected amino acid (0.5 mmol) in 4:1 DCM/DMF (4 mL) was treated with HOBt (0.6 mmol) and
HBTU (0.6 mmol), at r.t. and under inert atmosphere. After 5 min, serine ester (0.55 mmol), and
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DIPEA (1.2 mmol) were added and the reaction was stirred for 10 min under MW irradiation,
setting the internal reaction temperature at 80 °C. After concentration of the mixture at reduced
pressure, the residue was diluted with EtOAc (20 mL). The organic layer was washed with 0.1 M
HCl (5 mL), and a saturated solution of NaHCO3 (5 mL), and was dried over Na2SO4. After
evaporation of the solvent at reduced pressure, the crude dipeptide esters 3 were isolated (70-85 %,
80-90 % pure according to analytical RP HPLC) by flash chromatography over silica gel (eluent 1:3
cyclohexane/EtOAc), and were identified by ESI MS.
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Synthesis of N1-substituted Imi-peptides 7. General Procedure. To a solution of
triphenylphosphine (0.080 g, 0.31 mmol) in anhydrous DCM (4 ml), iodine (0.080 g, 0.32 mmol)
was added at rt under inert atmosphere and magnetic stirring. After 15 min, imidazole (0.043 g, 0.6
mmol) was also added and the mixture was stirred for additional 15 min. The dipeptide 3 (0.25
mmol), dissolved in DCM (2 ml), was finally added and the reaction mixture refluxed for 2 h. The
mixture was then cooled, diluted with DCM (6 ml), and washed with 10 % Na2S2O4 in water (4 ml).
The organic layer was dried over Na2SO4, and the solvents were evaporated at reduced pressure to
the final volume of 3 mL. The solution was allowed to stand at 5 °C overnight, and some white
solid Ph3PO which precipitated not quantitatively was removed by filtration and washed with DCM
(5 mL). The filtrate and the washes were evaporated at reduced pressure to give the crude iodides 4.
The crude iodides 4 (0.25 mmol) were dissolved in anhydrous DMF (5 mL), the amine (0.5
mmol) was added, and the mixture was heated at 50°C for 3 h. Then DMF was distilled at reduced
pressure, the residue was suspended in 0.1 M HCl (12 mL), and the mixture was washed with Et2O
(5 mL). The pH of the water layer was corrected to 9-10 with sat. Na2CO3, then it was extracted 3
times with EtOAc (5 mL). The collected organic layers were dried over Na2SO4, and solvent was
removed at reduced pressure. The crude residues were utilized for the next step without further
purifications, the only exception being 5d, which was purified as described thereafter.
Boc-Ala-Dap(allyl)-OMe (5d). The residue was purified by flash chromatography over silica gel
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(eluent EtOAc/MeOH 98:2), giving the dipeptide (0.058 g, 70 %, 94 % pure by RP HPLC). H
NMR (CDCl3, 400 MHz) 2 conformers  1.39 (d, J = 6.8 Hz, 3H, AlaMe), 1.44+1.45 (s, 9H, tBu),
1.69 (br. s, 1H, NH), 2.98 (dd, J = 4.4, 12.7 Hz, 1H, DapH), 3.08 (dd, J = 5.4, 12.7 Hz, 1H,
DapH), 3.22-3.27 (m, 2H, NCH2), 3.76 (s, 3H, OMe), 4.19 (m, 1H, AlaH), 4.63 (m, 1H,
DapH), 4.99 (br.d, 1H, AlaNH), 5.05-5.19 (m, 2H, =CH2), 5.82 (m, 1H, CH=), 6.92+7.06 (br.d,
1H, DapNH); 13C{1H} NMR (CDCl3, 100 MHz) 2 conformers  18.7, 28.6, 49.8, 52.3, 52.6, 52.8,
55.2, 77.6, 116.7+116.8, 136.5+136.6, 155.7, 172.0+172.1, 172.9+173.0; HRMS (ESI/QTOF) m/z:
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[M + H]+ Calcd for C15H28N3O5 330.2029; Found 330.2060. Anal. Calcd for C15H27N3O5: C,
54.70; H, 8.26; N, 12.76. Found: C, 54.84; H, 8.33; N, 12.87.
6
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The oily residues containing the crude dipeptides 5 were dissolved in DMF (5 mL), pNPC (0.075 g,
0.38 mmol) and DBU (0.055 g, 0.38 mmol) were added, and the mixture was stirred at rt for 12 h.
The solvent was distilled at reduced pressure, and 0.1 M HCl (4 mL) was added, then the residue
was extracted 3 times with EtOAc (5 mL). The collected organic layers were dried over Na2SO4,
and the solvent was removed at reduced pressure. The Imi dipeptides 7 were isolated from the
resulting mixtures by flash chromatography over silica gel (eluent EtOAc/MeOH 98:2), and purity
was determined by RP HPLC (General Methods).
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Boc-Ala-(S)-Imi(N1Me)-OMe (7a). Starting from 3a (0.070 g, 0.24 mmol), the General Procedure
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gave 7a (0.071 g, 90 %, 95 % pure). H NMR (CDCl3, 400 MHz)  1.38-1.48 (m, 12H,
AlaMe+tBu), 2.87 (s, 3H, NMe), 3.38 (dd, J = 3.4, 9.8 Hz, 1H, ImiH5), 3.71 (t, J = 10.0 Hz, 1H,
ImiH5), 3.77 (s, 3H, OMe), 4.82 (dd, J = 3.4, 9.8 Hz, 1H, ImiH4), 5.08 (d, J = 7.6 Hz, 1H, AlaNH),
5.51 (m, 1H, AlaH); 13C{1H} NMR (CDCl3, 100 MHz)  18.4, 28.3, 30.5, 46.6, 48.7, 51.9, 52.9,
79.6, 153.1, 155.3, 169.9, 174.1; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for C14H24N3O6
330.1665; Found 330.1688. Anal. Calcd for C14H23N3O6: C, 51.06; H, 7.04; N, 12.76. Found: C,
50.84; H, 7.28; N, 13.00.
Cbz-Val-(S)-Imi(N1iPr)-OMe (7b). Starting from 3b (0.088 g, 0.25 mmol), the General Procedure
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gave 7b (0.089 g, 85 %, 91 % pure). H NMR (CDCl3, 400 MHz)  0.86 (d, J = 7.2 Hz, 3H,
ValMe), 0.95 (d, J = 7.2 Hz, 3H, ValMe), 1.02 (d, J = 6.4 Hz, 3H, iPrMe), 1.10 (d, J = 6.4 Hz, 3H,
iPrMe), 2.25 (m, 1H, ValH), 2.35 (m, 1H, N1CH), 3.33 (dd, J = 3.6, 9.8 Hz, 1H, ImiH5), 3.66 (t, J
= 9.8 Hz, 1H, ImiH5), 3.78 (s, 3H, OMe), 4.82 (dd, J = 3.6, 9.8 Hz, 1H, ImiH4), 5.11 (br.s, 2H,
PhCH2), 5.41 (d, J = 8.0 Hz, 1H, ValNH), 5.61 (m, 1H, ValH), 7.33-7.41 (m, 5H, ArH); 13C{1H}
NMR (CDCl3, 100 MHz)  16.6, 18.8, 21.1, 22.6, 28.7, 41.4, 48.3, 50.2, 52.5, 56.7, 61.2, 128.0,
129.0, 129.1, 137.7, 153.4, 155.9, 173.5, 175.3; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for
C21H30N3O6 420.2135; Found 420.2172. Anal. Calcd for C21H29N3O6: C, 60.13; H, 6.97. N,
10.02. Found: C, 60.22; H, 7.26; N, 10.10.
Boc-Ala-(S)-Imi(N1Ph)-OBn (7c). Starting from 3c (0.084 g, 0.23 mmol), the General Procedure
1
gave 7c (0.072 g, 80 %, 92 % pure). H NMR (CDCl3, 400 MHz)  1.40 (d, J = 6.8 Hz, 3H,
AlaMe), 1.44 (s, 9H, tBu), 3.84 (dd, J = 3.2, 9.6 Hz, 1H, ImiH5), 4.19 (dd, J = 9.6, 10.0 Hz, 1H,
ImiH5), 5.00 (dd, J = 3.2, 10.0 Hz, 1H, ImiH4), 5.10 (d, J = 7.2, 1H, AlaNH), 5.17 (d, J = 12.0 Hz,
1H, PhCH), 5.26 (d, J = 12.0 Hz, 1H, PhCH), 5.53 (m, 1H, AlaH), 7.18 (m, 1H, ArH), 7.30-7.41
(m, 7H, ArH), 7.43-7.50 (m, 2H, ArH); 13C{1H} NMR (CDCl3, 100 MHz)  18.2, 28.3, 45.3, 49.1,
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51.6, 67.9, 79.8, 119.3, 124.9, 128.5, 128.7, 128.8, 129.1, 134.6, 137.9, 150.9, 155.3, 168.9, 174.4;
HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for C19H26N3O6 392.1822; Found 392.1856. Anal.
Calcd for C19H25N3O6: C, 58.30; H, 6.44; N, 10.74. Found: C, 58.47; H, 6.35; N, 10.83.
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Boc-Ala-(S)-Imi(N1allyl)-OBn (7e). Starting from 3c46 (0.091 g, 0.25 mmol), the General
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Procedure gave 7e (0.099 g, 93 %, 94 % pure). H NMR (CDCl3, 400 MHz)  1.36 (d, J = 7.2 Hz,
3H, AlaMe), 1,43 (s, 9H, tBu), 3.32 (dd, J = 3.0, 9.8 Hz, 1H, ImiH5), 3.65 (dd, J = 9.8, 10.4 Hz, 1H,
ImiH5), 3.78 (dd, J= 5.2, 14.8 Hz, 1H, CH2-CH=), 3.92 (dd, J= 6.2, 15.8 Hz, 1H, CH2-CH=), 4.87
(dd, J = 3.0, 10.4 Hz, 1H, ImiH4), 5.08 (d, J = 8.0 Hz, 1H, AlaNH), 5.17-5.27 (m, 4H,
=CH2+PhCH2), 5.49 (m, 1H, AlaH), 5.65 (m, 1H, -CH=), 7.30-7.40 (m, 5H, ArH); 13C{1H}
NMR, (CDCl3, 100 MHz)  18.7, 28.6, 44.5, 46.5, 49.2, 52.5, 68.1, 80.1, 119.5, 126.5, 128.8,
129.0, 131.5, 135.1, 153.1, 155.7, 169.6, 174.5; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for
C22H30N3O6 432.2135; Found 432.2166. Anal. Calcd for C22H29N3O6: C, 61.24; H, 6.77; N,
9.74. Found: C, 61.38; H, 6.84; N, 9.81.
Boc-(S)-Ala-(R)-Imi(N1Me)-OMe (7f). Starting from 3f (0.073 g, 0.25 mmol), the General
Procedure gave 7f (0.072 g, 87 %, 93 % pure). 1H NMR (CDCl3, 400 MHz)  1.36 (d, J = 7.2, 1H,
AlaMe), 1.42 (s, 9H, tBu), 2.88 (s, 3H, NMe), 3.38 (dd, J = 3.4, 9.7 Hz, 1H, ImiH5), 3.71 (dd, J =
9.7, 10.4 Hz, 1H, ImiH5), 3.76 (s, 3H, OMe), 4.70 (dd, J = 3.4, 10.4 Hz, 1H, ImiH4), 5.36 (br.d,
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1H, AlaNH), 5.51 (m, 1H, AlaH); C{1H} NMR (CDCl3, 100 MHz)  19.3, 28.2, 30.5, 46.6,
51.8, 52.4, 52.8, 79.3, 153.1, 154.8, 169.7, 173.9; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for
C14H24N3O6 330.1665; Found 330.1692. Anal. Calcd for C14H23N3O6: C, 51.06; H, 7.04; N,
12.76. Found: C, 50.76; H, 7.39; N, 13.04.
Cbz-Val-(R)-Imi(N1iPr)-OMe (7g). Starting from 3g (0.084 g, 0.24 mmol), the General Procedure
1
gave 7g (0.082 g, 82 %, 91 % pure). H NMR (CDCl3, 400 MHz)  0.83 (d, J = 7.2 Hz, 3H,
ValMe), 0.86 (d, J = 7.2 Hz, 3H, ValMe), 1.05 (d, J = 6.4 Hz, 3H, iPrMe), 1.10 (d, J = 6.4 Hz, 3H,
iPrMe), 2.19 (m, 1H, ValH), 2.37 (m, 1H, N1CH), 3.32 (dd, J = 3.6, 9.8 Hz, 1H, ImiH5), 3.66 (t, J
= 9.8 Hz, 1H, ImiH5), 3.78 (s, 3H, OMe), 4.69 (dd, J = 3.6, 9.8 Hz, 1H, ImiH4), 5.08-5.15 (m, 2H,
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PhCH2), 5.57-5.62 (m, 2H, ValH+ValNH), 7.28-7.39 (m, 5H, ArH); C{1H} NMR (CDCl3, 100
MHz)  16.3, 17.4, 20.3, 28.8, 40.5, 52.2, 53.4, 55.0, 59.2, 67.0, 127.9, 128.7, 129.1, 129.5, 137.1,
152.5, 155.0, 172.3, 174.5; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for C21H30N3O6 420.2135;
Found 420.2166. Anal. Calcd for C21H29N3O6: C, 60.13; H, 6.97; N, 10.02. Found: C, 60.30; H,
7.20; N, 10.19.
Boc-Ala-(R)-Imi(N1Ph)-OBn (7h). Starting from 3h (0.091 g, 0.25 mmol), the General Procedure
1
gave 7c (0.074 g, 77 %, 96 % pure). H NMR (CDCl3, 400 MHz)  1.44 (d, J = 6.4 Hz, 3H,
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AlaMe), 1.47 (s, 9H, tBu), 3.77 (dd, J = 2.6, 10.2 Hz, 1H, ImiH5), 4.23 (dd, J = 9.8, 10.2 Hz, 1H,
ImiH5), 4.88 (dd, J = 2.6, 9.8 Hz, 1H, ImiH4), 5.23 (s, 2H, PhCH2), 5.44 (br.d, 1H, AlaNH), 5.65
(m, 1H, AlaH), 7.19 (m, 1H, ArH), 7.30-7.41 (m, 7H, ArH), 7.43-7.51 (m, 2H, ArH); 13C{1H}
NMR (CDCl3, 100 MHz)  19.5, 28.4, 45.3, 49.5, 52.1, 67.9, 79.6, 119.2, 125.0, 126.1, 128.3,
128.7, 129.2, 134.8, 137.8, 150.8, 154.8, 168.7, 174.2; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd
for C19H26N3O6 392.1822; Found 392.1861. Anal. Calcd for C19H25N3O6: C, 58.30; H, 6.44; N,
10.74. Found: C, 58.44; H, 6.51; N, 10.90.
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Boc-Ala-(R)-Imi(N1allyl)-OBn (7i). Starting from 3h46 (0.088 g, 0.24 mmol), the General
Procedure gave 7e (0.093 g, 90 %, 92 % pure). 1H NMR (CDCl3, 400 MHz)  1.40 (d, J = 6.8 Hz,
3H, AlaMe), 1.46 (s, 9H, tBu), 3.29 (dd, J = 3.4, 9.8 Hz, 1H, ImiH5), 3.64 (dd, J = 9.8, 10.4 Hz, 1H,
ImiH5), 3.77 (dd, J = 5.8, 15.6 Hz, 1H, CH2-CH=), 3.96 (dd, J = 5.4, 15.6 Hz, 1H, CH2-CH=), 4.74
(dd, J = 3.4, 10.4 Hz, 1H, ImiH4), 5.15-5.22 (m, 4H, PhCH2+=CH2), 5.40 (br.d, 1H, AlaNH), 5.56
(m, 1H, AlaH), 5.66 (m, 1H, =CH2), 7.30-7.39 (m, 5H, Ar); 13C{1H} NMR (CDCl3, 100 MHz) 
19.9, 28.7, 44.4, 46.5, 49.5, 53.1, 68.0, 79.9, 119.4, 128.8, 128.9, 120.0, 131.5, 135.3, 152.9, 155.1,
169.4, 174.3; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for C22H30N3O6 432.2135; Found
432.2167. Anal. Calcd for C22H29N3O6: C, 61.24; H, 6.77; N, 9.74. Found: C, 61.33; H, 6.68; N,
9.66.
Boc-Ala-(S)-Imi[N1(CH2)3CO2Me]-OBn (7l). Starting from 3c46 (0.081 g, 0.22 mmol), the General
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Procedure gave 7l (0.080 g, 74 %, 94 % pure). H NMR (CDCl3, 400 MHz)  1.35 (d, J = 6.4Hz,
3H, AlaMe), 1.43 (s, 9H, tBu), 1.78-1.86 (m, 2H, CH2), 2.30-2.38 (m, 2H, CH2), 3.24 (m, 1H, CH2),
3.35-3.40 (m, 2H, CH2+ImiH5), 3.63-3.77 (m, 4H, COOMe+ImiH5), 4,87 (dd, J = 3.4, 10.2, 1H,
ImiH4), 5.07 (d, J = 8.4 Hz, 1H, AlaNH), 5.15 (d, J = 12.2 Hz, 1H, PhCH), 5.25 (d, J = 12.2 Hz,
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1H, PhCH), 5.46 (m, 1H, AlaH), 7.25-7.38 (m, 5H, ArH); C{1H} NMR (CDCl3, 100 MHz) 
14.6, 22.6, 28.7, 31.3, 43.4, 44.9, 48.0, 52.5, 68.2, 80.0, 128.8, 129.1, 143.1, 153.4, 155.9, 173.5,
175.3; HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for C24H34N3O8 492.2346; Found 492.2329.
Anal. Calcd for C24H33N3O8: C, 58.64; H, 6.77; N, 8.55. Found: C, 58.78; H, 6.82; N, 8.60.
(S)- or (R)-methyl 1-methyl-2-oxoimidazolidine-4-carboxylates (S)-8 or (R)-8. To a stirred
suspension of LiOH (0.028 g, 1.2 mmol) in THF (6 mL) and water (1.5 mL), a solution of H2O2 (30
wt %, 0.28 mL, 2.3 mmol) was added at 0 °C. After 15 min, a solution of 7a or 7f (0.13 g, 0.4
mmol) in THF (3 mL) was added at 0 °C while stirring. The temperature was allowed to raise to rt,
and after 4 h the pH was adjusted to 5-6 with 0.1 M HCl, THF was evaporated at reduced pressure,
and the mixture was extracted three times with EtOAc. The collected organic layers were dried over
Na2SO4, and the residue was purified by flash chromatography over silica gel (eluent
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cyclohexane/EtOAc 1:3) giving (S)-8 (0.050 g, 0.32 mmol, 80 % yield, 94 % pure) or (R)-8 (0.047
g, 0.30 mmol, 75 % yield, 94 % pure), their []D specific optical rotations were determined to be
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+25 (c 0.7, MeOH) and -24 (c 0.6, MeOH), respectively. Literature for (S)-8: []D +26.9 (c 1,
MeOH).23
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Boc-Ala-(R)-Imi(N1nPr)-Phe-OMe (9). The dipeptide 7i (0.1 g, 0.23 mmol) was subjected to
hydrogenation in the presence of a catalytic amount of 10 % Pd/C in EtOH (10 ML) at rt. The
reaction was monitored by TLC, and after 6 h the solution was filtered over celite®. The solvent
was removed at reduced pressure, and the presence of the dipeptide acid was confirmed by RP
HPLC and ESI MS.
The residue was dissolved in 3:1 DCM/DMF (6 mL), and reacted with HOBt (0.038 g, 0.28
.
.
mmol), H-Phe-OMe HCl (0.060 g, 0.28 mmol), and EDCI HCl (0.054 g, 0.28 mmol),
trimethylamine (0.083 mL, 0.60 mmol). The reaction was conducted as described for peptides 1,
and after the usual work up, the Imi-tripeptide 9 was isolated as described for the Imi-peptides 2
1
(0.090 g, 78 %, 88 % pure according to analytical RP HPLC, General Methods). H NMR (8:2
[D6]DMSO/H2O, 400 MHz)  0.74 (t, J = 7.3 Hz, 3H, Me), 1.16 (d, J = 6.4 Hz, 3H, AlaMe), 1.27-
1.34 (m, 2H, CH2), 1.36 (s, 9H, tBu), 2.56 (dd, J = 3.0, 9.5 Hz, 1H, ImiH5), 2.85 (dd, J = 10.8, 13.2
Hz, PheH), 2.95 (m, 1H, NCH2), 3.07-3.17 (m, 2H, PheH+NCH2), 3.46 (dd, J = 9.5, 10.0 Hz, 1H,
ImiH5), 3.65 (s, 3H, OMe), 4.50-4.60 (m, 2H, PheH+ImiH4), 5.34 (m, 1H, AlaH), 6.76 (d, J =
8.8 Hz, 1H, AlaNH), 7.17-7.22 (m, 3H, PheArH), 7.22-7.30 (m, 2H, PheArH), 8.59 (d, J = 8.0 Hz,
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1H, PheNH); C{1H} NMR ([D6]DMSO, 100 MHz)  11.7, 19.9, 20.5, 29.1, 38.1, 45.0, 45.7,
49.0, 53.0, 53.5, HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for C25H37N4O7 505.2662; Found
505.2685. Anal. Calcd for C25H36N4O7: C, 59.51; H, 7.19; N, 11.10. Found: C, 59.71; H, 7.22; N,
11.05.
Boc-Ala-(S)-Imi((E)-3-(4-((tert-butoxycarbonyl)amino) phenyl)allyl)-OBn (10). A mixture of
dipeptide 7e (0.056 g, 0.13 mmol), N-Boc-4-bromoaniline (0.054 g, 0.2 mmol), palladium acetate
(0.3 mg, 0.0013 mmol) and triphenylphosphine (0.7 mg, 0.0026 mmol) were stirred in 1:2
triethylamine/DMF (3 mL), under inert atmosphere. The mixture was stirred for 18 h at 80°C. Then
the mixture was evaporated at reduced pressure, MeOH (15 mL) was added to the residue and the
suspension was filtered through a Celite® pad to remove the fine black Pd°. The solvent was
evaporated at reduced pressure, the residue was dissolved in EtOAc (20 mL) and washed with 1M
HCl (4 mL). The organic layer was dried over Na2SO4 and after evaporation at reduced pressure the
residue was purified by semi-preparative RP HPLC over a C18 column (General Methods), giving
1
10 (0.063 g, 0.10 mmol, 78 %, 92 % pure by analytical RP HPLC, General Methods). H NMR
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(CDCl3, 400 MHz)  1.26 (s, 9H, tBu), 1.38 (d, J = 6.8 Hz, 3H, AlaMe), 1.44 (s, 9H, tBu), 3.35 (dd,
J = 3.4, 9.9 Hz, 1H, ImiH5), 3.70 (dd, J = 9.9, 10.3 Hz, 1H, ImiH5), 3.97 (dd, J = 6.4, 15.4 Hz, 1H,
CH-CH=), 4.07 (dd, J = 6.6, 15.4 Hz, 1H, CH-CH=), 4.88 (dd, J = 3.4, 10.3 Hz, 1H, ImiH4), 5.07
(d, J = 7.6 Hz, 1H, AlaNH), 5.13 (d, J = 12.2 Hz, 1H, CHPh), 5.23 (d, J = 12.2 Hz, 1H, CHPh), 5.51
(m, 1H, AlaH), 6.06 (m, 1H, CH=), 6.54 (d, J = 15.6 Hz, 1H, CH=), 7.28-7.36 (m, 7H, ArH), 7.39
(d, J = 8.8 Hz, 2H, ArH); 13C{1H} NMR (CDCl3, 100 MHz)  17.2, 28.7x2, 43.6, 48.5, 50.8, 53.1,
66.3, 81.7, 120.4, 124.0, 126.9, 129.0, 130.1, 132.2, 135.0, 137.8, 151.8, 152.8, 155.0, 172.1, 175.5;
HRMS (ESI/QTOF) m/z: [M + H]+ Calcd for C33H43N4O8 623.3081; Found 623.3101. Anal.
Calcd for C33H42N4O8: C, 63.65; H, 6.80; N, 9.00. Found: C, 61.41; H, 6.68; N, 9.05.
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Boc-Ala-(S)-Imi(3-(4-((tert-butoxycarbonyl)amino)phenyl) propyl)-OBn (11). The dipeptide 10
(0.063 g, 0.10 mmol) was treated with H2 in the presence of a catalytic amount of 10 % Pd/C in
EtOH at rt. The reaction was monitored by TLC, and after 8 h the solution was filtered over celite®,
and the solvent was removed at reduced pressure, giving dipeptide acid 11 (0.053 g, 99 %, 85 %
pure by analytical RP HPLC, General Methods). 1H NMR (CDCl3, 400 MHz)  1.12-1.23 (m, 12H,
tBu+AlaMe), 1.31, (s, 9H, tBu), 1.57-1.64 (m, 2H, CH2), 2.37-2.42 (m, 2H, CH2), 2.93-3.17 (m,
2H, CH2), 3.27 (m, 1H, ImiH5), 3.41 (m, 1H, ImiH5), 4.28 (m, 1H, ImiH4), 5.27 (m, 1H, AlaH),
5.66 (br.d, 1H, AlaNH), 6.96 (d, J = 8.0 Hz, 2H, ArH), 7.22 (d, J = 8.0 Hz, 2H, ArH), 10.50 (br.s,
13
1H, COOH); C{1H} NMR (CDCl3, 100 MHz)  17.5, 25.7, 27.1, 28.9, 31.7, 40.9, 48.0, 50.0,
55.8, 80.4, 81.0, 119.9, 121.1, 126.3, 127.6, 134.1, 135.3, 152.0, 153.0, 155.1, 171.9, 174.3; HRMS
(ESI/QTOF) m/z: [M + H]+ Calcd for C26H39N4O8 535.2768; Found 535.2804. Anal. Calcd for
C26H38N4O8: C, 58.41; H, 7.16; N, 10.48. Found: C, 58.66; H, 7.30; N, 10.20.
Acknowledgements
We thank MIUR (PRIN20157WW5EH) and Fondazione del Monte di Bologna e Ravenna
(IntegrAl-328bis/2017) for financial support.
Supporting Information
1
Tables S1, amide-NH temperature gradients; Tables S2-5, ROESY cross peaks for 2b-e; H and
13C{1H} NMR spectra.
References
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(1) a) Hruby, V. J.; Balse, P. M. Conformational and topographical considerations in designing
agonist peptidomimetics from peptide leads. Curr. Med. Chem. 2000, 7, 945-970; b) Abell, A. D.
Heterocyclic-based peptidomimetics. Lett. Pept. Sci. 2002, 8, 267-272; c) Gentilucci, L.; De Marco,
R.; Cerisoli, L. Chemical modifications designed to improve peptide stability: incorporation of non-
natural amino acids, pseudo-peptide bonds, and cyclization. Curr. Pharm. Des. 2010, 16, 3185-
3203; d) Liskamp, R. M. J.; Rijkers, D. T. S.; Kruijtzer, J. A. W.; Kemmink, J. Peptides and
proteins as a continuing exciting source of inspiration for peptidomimetics. ChemBioChem. 2011,
12, 1626-1653; e) De Marco, R.; Mazzotti, G.; Greco, A.; Gentilucci, L. Heterocyclic scaffolds in
the design of peptidomimetic integrin ligands: synthetic strategies, structural aspects, and biological
activity. Curr. Top. Med. Chem. 2016, 16, 343-359.
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35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(2) a) Ritchie, T. J. S. J. F.; Young, J.; Pickett, S. D. The impact of aromatic ring count on
compound developability: further insights by examining carbo- and hetero-aromatic and -aliphatic
ring types. Drug Discov. Today 2011, 16, 164-171; b) Lovering, F.; Bikker, J.; Humblet, C. Escape
from flatland: increasing saturation as an approach to improving clinical success. J. Med. Chem.
2009, 52, 6752-6756; c) Feher, M.; Schmidt, J. M. Property distributions:ꢀ differences between
drugs, natural products, and molecules from combinatorial chemistry. J. Chem. Inf. Comput. Sci.
2002, 43, 218-227; e) Gentilucci, L.; Gallo, F.; Meloni, F.; Mastandrea, M.; Del Secco, B.; De
Marco, R. Controlling cyclopeptide backbone conformation with β/α-hybrid scaffolds. Eur. J. Org.
Chem. 2016, 19, 3243-3251.
(3) (a) Keller, M.; Boissard, C.; Patiny, L.; Chung, N. N.; Lemieux, C.; Mutter M.; Schiller, P. W.
Pseudoproline-containing analogues of morphiceptin and endomorphin-2: evidence for a cis Tyr-
Pro amide bond in the bioactive conformation. J. Med. Chem. 2001, 44, 3896-3903; b) Gentilucci,
L.; Tolomelli, A.; De Marco, R.; Tomasini, C.; Feddersen, S. Synthesis of constrained
peptidomimetics containing 2‐oxo‐1,3‐oxazolidine‐4‐carboxylic acids. Eur. J. Org. Chem. 2011, 25,
4925-4930; c) De Marco, R.; Tolomelli, A.; Campitiello, M.; Rubini, P.; Gentilucci, L. Expedient
synthesis of pseudo-Pro-containing peptides: towards constrained peptidomimetics and foldamers.
Org. Biomol. Chem. 2012, 10, 2307-2317; d) De Marco, R.; Greco, A.; Rupiani, S.; Tolomelli, A.;
Tomasini, C.; Pieraccini, S.; Gentilucci, L. In-peptide synthesis of di-oxazolidinone and
dehydroamino acid-oxazolidinone motifs as β-turn inducers. Org. Biomol. Chem. 2013, 11, 4316-
4326; e) Galletti, P.; Soldati, R.; Pori, M.; Durso, M.; Tolomelli, A.; Gentilucci, L.; Dattoli, S. D.;
Baiula, M.; Spampinato, S.; Giacomini, D. Targeting integrins αvβ3 and α5β1 with new β-lactam
derivatives. Eur. J. Med. Chem. 2014, 83, 284-293; f) Tolomelli, A.; Baiula, M.; Viola, A.;
Ferrazzano, L.; Gentilucci, L.; Dattoli, S. D.; Spampinato, S.; Juaristi, E.; Escudero, M. Dehydro-β-
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The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
proline containing α4β1 integrin antagonists: stereochemical recognition in ligand-receptor interplay.
ACS Med. Chem. Lett. 2015, 6, 701-706; g) Dattoli, S. D.; Baiula, M.; De Marco, R.; Bedini, A.;
Anselmi, M.; Gentilucci, L.; Spampinato, S. DS-70, a novel and potent 4 integrin antagonist, is an
effective treatment for experimental allergic conjunctivitis in guinea pigs. Br. J. Pharmacol. 2018,
175, 3891-3910.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(4) a) Freidinger, R. M.; Veber, D. F.; Perlow, D. S.; Brooks, J. R.; Saperstein, R. Bioactive
conformation of luteinizing hormone-releasing hormone: evidence from a conformationally
constrained analog. Science 1980, 210, 656-658; b) Freidinger, R. M.; Perlow, D. S.; Veber, D. F.
Protected lactam-bridged dipeptides for use as conformational constraints in peptides. J. Org.
Chem. 1982, 47, 104-109; c) Freidinger, R. M. Synthesis of -lactam-constrained tryptophyl-lysine
derivatives. J. Org. Chem. 1985, 50, 3631-3635; d) Wolfe, M. S.; Dutta, D.; Aube, J.
Stereoselective synthesis of Freidinger lactams using oxaziridines derived from amino acids. J. Org.
Chem. 1997, 62, 654-663; e) Lee, H. J.; Song, J. W.; Choi, Y. S.; Park, H. M.; Lee, K. B. A
Theoretical study of conformational properties of N-methyl azapeptide derivatives. J. Am. Chem.
Soc. 2002, 124, 11881-11893; f) Jamieson, A. G.; Boutard, N.; Beauregard, K.; Bodas, M. S.; Ong,
H.; Quiniou, C.; Chemtob, S.; Lubell, W. D. Positional scanning for peptide secondary structure by
systematic solid-phase synthesis of amino lactam peptides. J. Am. Chem. Soc. 2009, 131, 7917-
7927; g) Boutard, N.; Jamieson, A. G.; Ong, H.; Lubell, W. D. Structure-activity analysis of the
growth hormone secretagogue GHRP‐6 by α‐ and β‐amino γ‐lactam positional scanning. Chem.
Biol. Drug Des. 2010, 75, 40-50; h) Greco, A.; Tani, S.; De Marco, R.; Gentilucci, L. Synthesis and
analysis of the conformational preferences of 5‐aminomethyloxazolidine‐2,4‐dione scaffolds: first
examples of β2‐ and β2,ꢀ2‐homo‐Freidinger lactam analogues. Chem.-Eur. J. 2014, 20, 13390-13404;
i) De Marco, R.; Mazzotti, G.; Dattoli, S. D.; Baiula, M.; Spampinato, S.; Greco, A.; Gentilucci, L.
5‐Aminomethyloxazolidine‐2,4‐dione hybrid α/β‐dipeptide scaffolds as inductors of constrained
conformations: applications to the synthesis of integrin antagonists. Biopolymers 2015, 104, 636-
649.
(5) a) Proulx, C.; Sabatino, D.; Hopewell, R.; Spiegel, J.; Ramos, Y. G.; Lubell, W. D. Azapeptides
and their therapeutic potential. Future Med. Chem. 2011, 3, 1139-1164; b) Proulx, C.; Lubell, W. D.
N-Amino-imidazolin-2-one peptide mimic synthesis and conformational analysis. Org. Lett. 2012,
14, 4552-4555; c) Proulx, C.; Lubell, W. D. Analysis of N-amino-imidazolin-2-one peptide turn
mimic 4-position substituent effects on conformation by X-ray crystallography. Biopolymers 2014,
102, 7-15; d) Skerlj, R.; Bridger, G.; Zhou, Y.; Bourque, E.; McEachern, E.; Metz, M.; Harwig, C.;
Li, T.; Yang, W.; Bogucki, D.; Zhu, Y.; Langille, J.; Veale, D.; Ba, T.; Bey, M.; Baird, I.;
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