1,5-Disubstituted 1,2,3-Triazole-Containing Peptidotriazolamers: Design Principles for a Class of Versatile Peptidomimetics

Article abstract of DOI:10.1002/chem.201704583

Peptidotriazolamers are hybrid foldamers combining features of peptides and triazolamers—repetitive peptidomimetic structures with triazoles replacing peptide bonds. We report on the synthesis of a new class of peptidomimetics, containing 1,5-disubstituted 1,2,3-triazoles in an alternating fashion with amide bonds and the analysis of their conformation in solid state and solution. Homo- or heterochiral peptidotriazolamers were obtained from enantiomerically pure propargylamines with stereogenic centers in the propargylic position and α-azido esters by ruthenium-catalyzed azide–alkyne cycloaddition (RuAAC) under microwave conditions in high yields. With such building blocks the peptidotriazolamers are readily available by solution phase synthesis. While the conformation of the homochiral peptidotriazolamer Boc-Ala[5Tz]Phe-Val[5Tz]Ala-Leu[5Tz]Val-OBzl resembles that of a β VIa1 turn, the heterochiral peptidotriazolamer Boc-d-Ala[5Tz]Phe-d-Val[5Tz]Ala-d-Leu[5Tz]Val-OBzl adopts a polyproline-like repetitive structure.

Full text of DOI:10.1002/chem.201704583

A Journal of
Accepted Article
Title: 1,5-Disubstituted 1,2,3-Triazole-Containing Peptidotriazolamers:
Design Principles for a Class of Versatile Peptidomimetics
Authors: Oliver Kracker, Jerzy Góra, Joanna Krzciuk-Gula, Antoine
Marion, Beate Neumann, Hans-Georg Stammler, Anke Nieß,
Iris Antes, Rafal Latajka, and Norbert Sewald
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To be cited as: Chem. Eur. J. 10.1002/chem.201704583
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1,5-Disubstituted 1,2,3-Triazole-Containing Peptidotriazolamers:
Design Principles for a Class of Versatile Peptidomimetics
Oliver Kracker,[a] Jerzy Góra,[a],[d] Joanna Krzciuk-Gula,[a],[d] Antoine Marion,[c] Beate Neumann,[b] Hans-
Georg Stammler,[b] Anke Nieß,[a] Iris Antes,[c] Rafał Latajka,[d] and Norbert Sewald*[a]
Abstract: Peptidotriazolamers are hybrid foldamers combining
features of peptides and triazolamers – repetitive peptidomimetic
structures with triazoles replacing peptide bonds. We report on the
synthesis of a new class of peptidomimetics, containing 1,5-disub-
stituted 1,2,3-triazoles in an alternating fashion with amide bonds
and the analysis of their conformation in solid state and solution.
Homo- or heterochiral peptidotriazolamers were obtained from enan-
tiomerically pure propargylamines with stereogenic centers in the
propargylic position and α-azido esters by ruthenium-catalyzed
azide-alkyne cycloaddition (RuAAC) under microwave conditions in
high yields. With such building blocks the peptidotriazolamers are
readily available by solution phase synthesis. While the conformation
of the homochiral peptidotriazolamer Boc-Ala[5Tz]Phe-Val[5Tz]Ala-
Leu[5Tz]Val-OBzl resembles that of a β VIa1 turn, the heterochiral
peptidotriazolamer Boc-D-Ala[5Tz]Phe-D-Val[5Tz]Ala-D-Leu[5Tz]Val-
OBzl adopts a polyproline-like repetitive structure.
of oligopyrrolinones as β-strand peptidomimetics. They contain a
strongly modified peptide backbone integrated in cyclic struc-
tures also featuring vinylogous amino acids (Figure 1).[4] Similar
oligomers composed of 1,4-disubstituted triazoles were de-
scribed by Arora et al. as being reminiscent of a peptide β-strand
conformation.[5- 7] These triazolamers are somewhat unique as
they connect a series of triazole rings by single C1 units that, at
the same time, is a stereogenic centre (Figure 1).
O
O
HN
φ
ω
ψ
Me
CO2
HN
N
HN
O
N
N
N
N
N
BocHN
N
N
N
Me
CO2
R
Introduction
Figure 1. Examples of peptidomimetics: oligopyrrolinones[4] (top) and 1,4-
disubstituted triazolamers (bottom).[5-7]
Discrete stable conformations are prerequisites of functional
biopolymers, e.g., proteins. Many unique functions of peptides
and proteins, including molecular recognition, rely on the preva-
lence of secondary and/or tertiary structures. Foldamers, a term
coined by Gellman,[1] are repetitive structures of naturally occur-
ring or synthetic building blocks that emerged as analogues or
surrogates of native biomolecules. They were in many cases
proven to be functional biomimetic devices.[2,3]
.
The synthesis of triazoles by thermal 1,3-dipolar cycloaddition of
alkynes and azides was first described by Michael et al.[8] and
later studied systematically by Huisgen.[9] The thermal reaction,
e.g., between phenylazide and phenylacetylene, gives a nearly
equimolar mixture of the 1,4- and 1,5-disubstituted diphenyltria-
zole. The copper(I) catalysed azide-alkyne cycloaddition (Cu-
AAC), which was described independently by the groups of
Meldal[ 10] and Sharpless[ 11] in 2002, exclusively leads to 1,4-
disubstituted 1,2,3-triazoles. The CuAAC can be performed
either in solution or on solid phase, and has found widespread
application.[12- 15] This is mainly due to the key features of this
versatile reaction: i.e., the reaction proceeds very selectively and
without side products in organic or in aqueous media, and is
orthogonal towards biologically relevant functional groups. Be-
cause of its perfect atom economy and straightforward workup,
Sharpless and co-workers considered the CuAAC as a prototype
of a “click-reaction”.[13] In 2005 its counterpart, the ruthenium(II)-
catalysed-azide alkyne-cycloaddition (RuAAC) was present-
ed.[ 16 ] In the presence of Cp*RuCl(L2) the 1,5-disubstituted
1,2,3-triazoles are formed. Although the RuAAC does not toler-
ate water or protic solvents, it is quite tolerant towards functional
groups like alcohols, aldehydes, alkenes, amides, amines, bo-
ronic esters, ketones and halides.[17]
Hirschmann and co-workers presented in 1992 the novel class
[a]
O. Kracker, J. Góra, Dr. J. Krzciuk-Gula, A. Nieß, Prof. Dr. N.
Sewald*
Organic and Bioorganic Chemistry, Department of Chemistry
Bielefeld University
PO Box 100131, D-33501 Bielefeld, Germany
E-mail: norbert.sewald@uni-bielefeld.de
[b]
[c]
[d]
B. Neumann, Dr. H.-G. Stammler
Inorganic and Structural Chemistry, Department of Chemistry
Bielefeld University
PO Box 100131, D-33501 Bielefeld, Germany
Dr. A. Marion, Prof. Dr. I. Antes
Center for Integrated Protein Science, Department of Biosciences
TU Munich
Emil-Erlenmeyer-Forum 8, D-85354 Freising, Germany
J. Góra, Dr. J. Krzciuk-Gula, Prof. Dr. R. Latajka
Department of Bioorganic Chemistry
Wrocław University of Science and Technology
Wybrzeze Wyspianskiego 27, PL-50-370 Wrocław, Poland
Triazoles are considered peptide bond surrogates in pep-
tidomimetics and to be suitable substitutes for amide bonds
Supporting information for this article is given via a link at the end of
the document.
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10.1002/chem.201704583
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(Figure 2).[18,19] Just like amide bonds, they are planar, have a
strong dipole moment and are capable of accepting H-bonds.
Additionally, they are proteolytically and metabolically stable.
While the 1,5-disubstituted triazoles closely resemble a cis-
configured peptide bond, the 1,4-disubstituted triazole does not
perfectly match a trans-configured peptide bond.
Swern oxidation.[25] However, these α-amino aldehydes are very
prone to epimerization during prolonged storage or upon contact
with silica and are, therefore, to be used immediately.
O
O
P
O
R
R
O
R
N2
reduction
OH
H
3.7
4.9
BocHN
BocHN
Å
Å
BocHN
R
R
K
2CO3,
O
O
R
R
RT
MeOH,
C
O
C
H
N
N
7
6
9
N
N
C
N
C
C
H
H
N
C
N
3.0
3.2
Å
N
N
R
Å
N
C
NH
H
H
N
O
R
R
C
R
CBr4, PPh3
RT
BuLi, THF
-78 °C
R
Br
DCM,
trans amide bond
cis amide bond
1,4-disubstituted
triazole
1,5-disubstituted
triazole
BocHN
Br
8
Scheme 2. Synthesis of chiral propargylamines by the Corey-Fuchs or Best-
mann-Ohira reaction.
Figure 2. Comparison between trans- and cis-peptide bonds and their triazole
analogues.[20]
While several protocols for the synthesis of chiral propargyl-
amines have been published, most of these publications are
solely comparing the optical rotation of the final product. Recent
publications reported partial epimerization upon reaction of the
Bestmann-Ohira reagent with chiral α-amino aldehydes.[ 26- 28]
The risk of epimerization at the aldehyde stage is increased
when electron withdrawing sidechain residues are present that
further increase the acidity in α-position to the carbonyl group.
Recrystallization of a partially epimerised N-protected chiral
propargylamine was shown to increase the enantiomeric ex-
cess.[29]
O
TFA
N
H2N
N
O
N
2
O
N
BocHN
Boc-(Gly
Boc-(Gly
Gly) -OMe
2
[5Tz]
4
N
O
T P NEt
/
3
3
N
1
repeat
deprotection
coupling
NaOH
OH
N
BocHN
N
Gly) -OMe
O
[5Tz]
5
4
N
3
Both methods rely on the natural chiral pool. In addition, robust
methodology for the synthesis of chiral propargylamines with
non-natural side chain residues is available with Ellman’s tert-
butylsulfinamide as a chiral auxiliary (Scheme 3).[30- 33] This auxi-
liary reacts with aldehydes in a condensation reaction to form N-
sufinyl aldimines 10, which are further converted by organome-
tallic reagents 11 to give chiral amines 12.
The N-sulfinyl group can be cleaved under acidic conditions to
release the free amine as a salt. Hence, the N-sulfinyl group has
been suggested to be used as a protecting group similar to the
Boc group. Moreover, after acidic cleavage, it was shown that
the auxiliary can be recovered from the reaction.[34,35]
Scheme 1. Building block approach for the synthesis of a glycine-based
peptidotriazolamer.
In 2014, Johansson et al. published the synthesis of achiral
peptidomimetic foldamers containing 1,5-disubstituted 1,2,3-
triazoles and amide bonds in an alternating fashion
(Scheme 1).[ 21 ] As the triazole is considered a peptide bond
surrogate, we suggest to adapt the peptidomimetics nomencla-
ture accordingly: If the amide bond -(C=O)-NH- in the dipep-
tide -Gly-Gly- is replaced by a 1,5-disubstituted triazole it is
represented by -Gly[5Tz]Gly-.
Because of the polarity of the triazole units and amide bonds,
these oligomers are water soluble with the solubility slightly
increasing with chain length. Conformational analysis of the
oligomers and quantum chemical calculations revealed the co-
existence of several conformers with comparable stabilities.[21]
Based on the fact that achiral monomers were used, the sec-
ondary structure motifs are independent on different side chains
and rather formed by backbone interactions.
Triazoles substituted with adjacent stereogenic centres require
enantiomerically pure propargylamines and α-azido acids. The
synthesis of chiral propargylamines 9 has been described using
protected α-amino aldehydes 7 in a Corey-Fuchs or Bestmann-
Ohira reaction (Scheme 2).[ 22- 24] The α-amino aldehydes are
accessible by either reduction of the corresponding Weinreb
amide, by LiAlH4/DIBAL-H, or oxidation of the amino alcohol by
O
S
R
O
S
+
ClMg
TMS
N
N
R
rt
to
H
-78
DCM,
°C
eq
TMS
2-3
10
11
12
Scheme 3. Diastereoselective synthesis of chiral N-sulfinyl propargylamines
i
(R= Pr, tBu, CH2OTBDPS), Ph, p-Cl-Ph, p-NO2-Ph, p-MeO-Ph, thienyl, furyl,
Boc-pyrrolyl.[30-33]
Foldamers composed from building blocks presenting amino
acid side chains and able to adopt a regular and predictable
conformation have potential for controlling protein-protein inter-
action and peptide aggregation.[1-3]
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10.1002/chem.201704583
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Results and Discussion
(Scheme 5).[36,37] By employing Wong’s procedure, the azides
22a-c were synthesized in excellent yields of >87%, starting
from commercially available amino acid benzyl ester salts 21.
We embarked on a bottom-up approach investigating the con-
formational behaviour of homo- (13) or heterochiral (14) pepti-
dotriazolamers as modulators of protein-protein interaction (Fig-
R
R
TfN3, K2CO3, CuSO4
X-
O
O
+H3N
N3
ure 3).
H
2O / DCM /
tBuOH
N
O
O
R
R
R
O
R
N
H
N
21
22
N
N
Bzl
CO2
N
N
H
BocHN
N
=
R
R
R
iPr
N
R
R
O
22a
22b
22c
(98%)
N
N
=
=
Me
Bzl
(93%)
(87%)
13
homochiral
Scheme 5. Synthesis of chiral α-azido esters.
N
N
N
R
R
R
O
R
H
N
N
Bzl
CO2
N
N
H
BocHN
N
N
R
R
O
N
N
The (R) or (S) configured propargylamines were obtained using
Ellman’s auxiliary as described by Wünsch et al.[33] Although the
tert-butylsulfinyl (Bus) group had been previously discussed as a
substitute for the Boc group in peptide synthesis, we found it to
be unstable under microwave conditions during the RuAAC.
Moreover, it decomposes upon standing in polar solvents. A
thermal rearrangement of the sulfinamides as investigated by
Arava et al.[38] was identified to be responsible for low yields in
the RuAAC. Accordingly, N-sulfinyl propargylamines react inter-
molecularly forming deprotected and N-(tert-butylthio)-tert-butyl-
sulfonamide linked alkyne. Because of this prominent side reac-
tion, the Bus-protected propargylamines 23 were deprotected
(24), and the Boc group was introduced (25) (Scheme 6).
14
heterochiral
Figure 3. Homochiral and heterochiral peptidotriazole hexamers.
Peptidotriazolamer building blocks of type 20 were synthesized
in solution with orthogonal Boc/Bzl protection using triazole
derivatives 17. The C-terminal ester is indispensable, since the
RuAAC is not compatible with free carboxylic acids (Scheme 4).
Such peptidotriazolamers can be synthesized according to a
building block approach using triazole derivatives previously
synthesized from a propargylamine and an α-azidocarboxylic
acid. Alternatively, a submonomer approach involving amide
bond formation of an α-azidocarboxylic acid, followed by RuAAC
with a protected propargylamine would be feasible.
O
eq)
eq)
1) (Boc)2 (2
NaHCO
3 (4
R
O
S
R
R
HCl
eq)
imidazole
(3
eq)
(2
2)
+H3N
Cl-
BocHN
N
H
MeOH
H
THF 1:1
2O /
23
24
25
+
=
=
=
=
=
=
25a R
25b R
25c R
25d R
25e R
25f R
R
R
R
R
(S)-Me (61%)
(R)-Me (85%)
OBzl
OBzl
RuAAC
BocHN
15
N3
16
N
BocHN
iPr
(S)- (56%)
N
O
O
iPr
(R)- (90%)
N
17
iBu
(S)-
(R)-
(82%)
(94%)
iBu
H
2, Pd/C
HCl
R
Scheme 6. Acidic methanolysis of the Bus group, followed by Boc protection
R
N
R
R
leads to Boc protected propargylamines in good yields over two steps.
OH
OBzl
N
BocHN
H2N
N
N
N
N
18
O
O
19
Based on a publication by Basel et al., imidazole was used to
scavenge the slight excess of (Boc)2O, which can be removed
by washing with dilute acid, thus avoiding the necessity of a
column chromatography in most cases.[39] Using this procedure,
the Boc protected (R) or (S) configured alkyne analogues of Ala
(25a,b), Val (25c,d), and Leu (25e,f) were successfully synthe-
sized in enantiomerically pure form.
peptide
coupling
N
R
O
R
N
H
N
N
N
OBzl
BocHN
N
R
R
O
N
20
The RuAAC was performed under microwave conditions. THF
turned out to be a suitable solvent, regarding solubility of the
starting material, boiling temperature and stability of the catalyst.
Noteworthy, additional degassing of freshly destilled THF was
not required. Using a slight excess of azide 22, the RuAAC was
complete in 1-2 h providing different homo- and heterochiral
triazoles 28-33 representing dipeptide isosters in excellent yields
of >80% (Scheme 7).
Scheme 4. Envisaged strategy for solution synthesis of peptidotriazolamers
using 1,5-disubstituted triazole building blocks obtained by RuAAC.
The synthesis of chiral α-azido esters 22 preferably makes use
of the pool of naturally occurring amino acids that can be con-
verted by Cu(II)-catalysed diazo transfer from triflyl azide
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free preactivation in DCM, instead of DMF.[40] The problem of
possible epimerization during the coupling of triazole dipeptide-
isosteric building blocks was also reported for 1,4-disubstituted
triazoles.[41,42] Horne et al. overcame this problem by coupling
with a five-fold excess of triazole/DIC/HOAt to a free amine on
solid phase, without the use of any base for the coupling step.
In our work 2,4,6-trimethylpyridine (sym-collidine) was used as a
base to liberate the amine from the hydrochloride salt, obtained
upon cleavage of the Boc group. The peptide coupling was not
accompanied by epimerization at the triazole substituted stereo-
genic centres when using a base-free, carbodiimide mediated
preactivation of the carboxylic acid, in combination with Oxyma-
Pure or HOAt, respectively.[43] Hence, a 1.1-fold excess of acti-
vated acid, in combination with sym-collidine as a base for liber-
ating the amine from the ammonium salt, was sufficient to full
conversion of the amine. Under these conditions, coupling pro-
ceeded in DCM without any epimerization and with high yields
(Schemes 8, 9).
R2
R1
R1
R2
Cp*RuCl(COD)
(3-5mol%)
+
N
Bzl
CO2
BocHN
BocHN
N3
Bzl
CO2
THF, 1-2 h
N
MW,
20W
60°C,
N
26
22
27
N
Bzl
CO2
BocHN
N
N
Bzl
CO2
N
Bzl
CO2
BocHN
BocHN
N
N
N
N
N
28
29
30
(88%)
(87%)
(94%)
N
Bzl
CO2
N
N
N
Bzl
CO2
BocHN
BocHN
N
N
N
Bzl
CO2
BocHN
N
N
31
32
33
(84%)
(81%)
(88%)
H
Pd/C
,
2
1)
2)
3)
4)
Scheme 7. Microwave-assisted RuAAC providing 1,5-disubstituted triazole
building blocks.
DIC/Oxyma
H-Leu
Val-OBzl
N
N
[5Tz]
collidine
H
N
N
Bzl
CO2
sym-
N
BocHN
Boc-Val
Ala-OBzl
[5Tz]
N
O
N
33
The crystallographic data confirm the formation of the 1,5-
disubstituted regioisomer in the RuAAC. Moreover, it highlights
the turn inducing effect of the 1,5-disubstituted triazole, as
shown for Boc-Val[5Tz]Ala-OBzl (33) as an example, with a
distance of 3.17 Å between both α-carbons, displaying close
resemblance to a cis-peptide bond (Figure 4) (see Supporting
Information for additional crystallographic data).
34
Val-OBzl
[5Tz]
Boc-Val
Ala-Leu
[5Tz]
(81%)
HCl
sym
1)
2)
H
,
Pd/C
DIC/Oxyma
1)
2
-collidine
2)
Boc-Ala
Phe-OBzl
[5Tz]
28
N
N
N
O
H
N
N
Bzl
CO2
N
N
BocHN
H
N
N
O
N
N
35
Val-OBzl
[5Tz]
Boc-Ala
Phe-Val
Ala-Leu
[5Tz]
[5Tz]
(84%)
Scheme 8. Synthesis of the hexameric homochiral peptidotriazole 35.
H
Pd / C
,
1)
2)
3)
4)
2
DCC / HOAt,
H- -Val
Ala-OBzl
D
[5Tz]
-collidine
N
sym
O
N
N
H
Boc- -Ala
Phe-OBzl
D
[5Tz]
Figure 4. Crystal structure of Boc-Val[5Tz]Ala-OBzl (33), displaying the chain
reversal induced by the 1,5-disubstituted triazole.
N
N
OBzl
BocHN
29
N
O
N
Boc- -Ala
Phe- -Val
36
Ala-OBzl
D
D
[5Tz]
[5Tz]
(51%)
With the homo- and heterochiral triazole dipeptide building
blocks 28-33 in hand, solution phase peptide synthesis was
used for the assembly of larger peptidotriazolamers. The acidic
cleavage of the Boc group and hydrogenolysis of the benzyl
ester, resp., were quantitative. The deprotected derivatives were
used without further purification for the peptide coupling
(Schemes 8, 9). With TBTU or HATU as coupling reagents in
DMF, either without or in combination with HOBt/HOAt and
DIPEA as a base, the products were obtained in high yields, but
suffered from epimerization. Carpino and El-Faham had demon-
strated the influence of solvent and base on the preactivation
rate of amino acids with DIC/HOAt and suggested to use a base-
H
Pd / C
,
1)
2)
3)
4)
2
DCC / HOAt,
H- -Leu
Val-OBzl
D
[5Tz]
-collidine
sym
N
O
N
H
O
N
N
N
N
N
N
H
BocHN
N
N
O
O
N
Boc- -Ala
Phe- -Val
Ala- -Leu
Val-OBzl
[5Tz]
D
D
D
[5Tz]
[5Tz]
(70%)
37
Scheme 9. Synthesis of the hexameric heterochiral peptidotriazole 37.
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The peptidotriazolamers could be purified by either column
chromatography or preparative HPLC. In contrast to Johansson
et al., [21] a drastic decrease in coupling yields of the tetramer to
give the hexamer units was not observed. Furthermore, the
carbodiimide mediated coupling provides increased coupling
yields, compared to the T3P/DIPEA system Johansson et al.
used for their achiral oligomers. If solubility of the hydrochloride
salt in DCM becomes a problem, it can be dissolved in DMF and
added dropwise to the preactivated acid, dissolved in DCM, for a
final DCM/DMF ratio of 1:1 without any epimerization.
Conformational analysis
The X-ray single crystal structure analysis of the tetramer Boc-
Val[5Tz]Ala-Leu[5Tz]Val-OBzl (34) revealed the torsion angles
of the peptide backbone in the solid state (Figure 5, Table 1).
The conformation is being stabilized by an intramolecular hydro-
gen bond between the Boc carbonyl oxygen and the amide
proton of Leu, with a distance of 2.08 Å.
We also modeled the structures of 34, 35, and 37 by means of
molecular dynamics (MD) simulations to further characterize the
conformational preferences of the 1,5-substituted peptidotriazol-
amers. The simulations were based on the specific TZLff molec-
ular mechanics force-field parametrization of 1,4- and 1,5-substi-
tuted triazole-based peptidomimetics that we recently devel-
oped.[ 44] As the backbone of peptidotriazolamers differs from
standard polypeptides, we use here the following nomenclature
to label the torsion angles: φn=C(c=0)-N(NH)-Cα-C5; ψn=N(NH)-Cα-
Figure 5. Comparison of solid-state (top, left) and simulated (top, right) con-
formation of the peptidotriazolamer 34 and an overlay of both structures
(bottom; all atoms RMSD = 0.37 Å; green: X-ray; magenta: MD).
C5-N1; φc=C5-N1-Cα-C(C=O); ψc=N1-Cα-C(C=O)-N(NH)
.
For peptidotriazolamers 35 and 37 the first stage of the calcula-
tions consisted of NMR data refinement by means of gas-phase
simulated annealing followed by an MD simulation in the original
NMR solvent (DMSO) with ROESY-derived distance restraints
imposed on both calculations. Furthermore, we performed addi-
tional unrestrained MD simulations in DMSO and water in order
to validate the adequacy of the initial set of distance restraints,
sample regions of the conformational space that may have been
previously inaccessible and observe the behavior of the ana-
lyzed compounds in a different solvent model.
During the distance-restrained MD simulation run the torsional
angles of 35 oscillated around the values produced by the SA
run (Figure 6A). This indicates that the initial set of NMR-derived
distance restraints manages to reproduce the conformational
behavior of 35 even without the presence of explicit solvent. At
300 K the system samples a restricted number of conformations,
similar to the final structure obtained from SA and has a fairly
compact structure (Figures 5A, 7A).
We performed a non-restrained simulated annealing (SA) of
peptidotriazolamer 34 in the gas phase to compare the structure
with the solid-state structure. The simulation was started from a
random conformation of the molecule, far from that of the crystal
structure, in order to avoid any bias in our modeling attempt. The
final structure is depicted in Figure 5 and the values of backbone
torsions are reported in Table 1, along with those of the X-ray
analysis. Both structures show a nearly identical backbone
conformation, with a bend in the middle of the sequence that is
stabilized by an intramolecular hydrogen bond. Further simula-
tions of 34 in DMSO are reported in our previous publication.[44]
The agreement between experimental and modeled structures is
very good, making us confident about the validity of the next
step of this study.
Table 1. Comparison of the torsion angles of the tetramer 34 according to X-
ray analysis and molecular dynamics (MD) simulation.
φn/c
ψn/c
Table 2. Torsion angles for the representative structure of 35 observed during
distance-restrained MD in DMSO and water.
X-ray
MD
X-ray
MD
Val1
Ala2
Leu3
Val4
-66.4(2)°
-112.2(2)°
-128.6(1)°
-93.2(2)°
-66.3°
132.8(1)°
39.8(2)°
57.7(2)°
130.9°
30.4°
57.1°
φn/c (DMSO)
-112.9°
-92.6°
ψn/c (DMSO)
62.8°
83.1°
123.9°
52.1°
99.2°
-
φn/c (water)
-137.2
-120.2
-79.7
ψn/c (water)
74.2
7.5
-111.4°
-131.0°
-100.4°
Ala1
Phe2
Val3
Ala4
Leu5
Val6
-61.1°
116.4
1.9
-108.5°
-91.3°
-100.0
-132.4
-50.0
136.2
-
-166.0°
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Figure 6. φ/ψ Distribution for 35 during distance-restrained MD in DMSO (A; black dots annotate torsional angle values from SA) and unrestrained simula-
tions in DMSO (B) and in water (C).
The representative conformation from clustering over the last
100 ns of the simulation resembles that of a β-turn for natural
peptides. It is stabilized by a well-conserved hydrogen bond
(persisting for 91.7% of the clustered trajectory) formed between
the i and i+3 residues (Phe2 and Leu5). The corresponding
values of torsional angles for the i+1 and i+2 residues (Table 2)
predominantly to the side chains and the Boc protecting group,
which was not involved in any distance restraint. As a result, two
additional basins at (-160°, 160°) and (160°, -160°) of the phi/psi
distribution plot can be observed in contrast to the data estab-
lished during SA.
This highlights the advantage of using an explicit solvent model
suggest that it resembles a type VIa1 β-turn. In this conformation, for the improvement of conformational space sampling, especial-
the backbone of the molecule is folded towards the inner side of
the turn, whereas the side chains are protruding towards the
solvent. The overall conformation of 35 is also in good agree-
ment with X-ray data obtained for its precursor 34 (Figure 4,
Table 1). The torsional angle values are comparable and both
compounds form a turn stabilized by an internal hydrogen bond.
The heterochiral peptidotriazolamer 37 displays more flexibility
upon sampling of the conformational space in course of the
distance-restrained MD simulation (Figure 7A, Table 3). This
was an expected result as a lower number of crosspeaks could
be observed on the ROESY spectra in comparison to 35. Ac-
cordingly, only a total of 22 distance restraints was generated for
37 in contrast to 55 restraints for 35. The peptidotriazolamer 37
displayed a tendency to adopt an extended conformation, which
allows accessing a wider variety of conformations. This applies
ly in case of extended conformations, where it is likely to ob-
serve less cross-peaks on ROESY and NOESY spectra. The
representative structure (Figure 9A) is not stabilized by any
intramolecular hydrogen bond. The triazole rings are located on
the same side of the backbone and are almost stacked upon
one another, while the side chains are presented on the oppo-
site site of the structure. The overall fold bears a distant similari-
ty to the polyproline family of helices. It resembles a left-handed
helix with no hydrogen bonding and three residues per turn,
which are the features of a polyproline helix. It is, however, more
denser and contains cis peptide bond analogues. Thus, it is
reminiscent of the polyproline I type helix.
Subsequently, we performed unrestrained simulations in DMSO
to assess a more realistic picture of the dynamics of the two
peptidomimetics. As expected, the release of the constraints
Figure 7. φ/ψ Distribution for 37 during distance-restrained MD in DMSO (A; black dots annotate torsional angle values from SA) and unrestrained simula-
tions in DMSO (B) and in water (C).
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resulted in sampling of new regions, which were inaccessible
during the previous simulations (Figures 5B, 7B). The effects are
more apparent for 37 (Figure 7B) because of its extended con-
formation and lack of intramolecular stabilization. For both com-
pounds the overall backbone conformation of their representa-
tive structures is very similar to the one derived from distance-
restrained MD. Two major clusters (45.9% and 25.9% of the
clustered population) can be distinguished for 35 (representative
structure of dominant cluster, Figure 8B; representatives of the
main clusters, Figure SI 23) and three (36.5%, 23.3% and 12.3%
of the clustered population) for 37 (representative structure of
dominant cluster, Figure 9B; representatives of the main clusters,
Figure SI 24). The two conformers of 35 share a similar fold of
the backbone with slight variations of the more flexible extremi-
ties. In case of 37 a larger fragment of the N-terminus, consist-
ing of the Boc protecting group and the first two amino acid
residues (Ala1, Phe2) exhibit an increased degree of freedom
and fold onto the backbone. In this scenario, there are three
distinct positions of the N-termini for the conformers of 37.
Therefore, when comparing restrained and conventional MD
simulations, an additional basin around (60°, 60°) appears on
the φ/ψ distribution plot of the latter (Figure 7B, Table 3). The
remaining residues of the molecule adopt a conformation that is
well preserved among the representatives of the three clusters.
Table 3. Torsion angles of the representative structure of 37 observed during
restrained MD in DMSO and water.
φn/c (DMSO)
145.6
-135.3
84.3
ψn/c (DMSO)
-140.9
151.8
-92.3
43.0
φn/c (water)
163.9
-130.3
102.4
-95.7
ψn/c (water)
-83.2
39.2
-87.9
39.4
-90.4
-
D-Ala1
Phe2
D-Val3
Ala4
-106.2
92.9
D-Leu5
Val6
-103.3
-
100.6
-68.6
-71.9
To gain insights into the behavior of the compounds in a more
biological environment, we performed unrestrained MD runs in
pure TIP4Pew water.[45] Changing the solvent polarity restricts
the conformational space for 35 and 37. Within the limits of the
current simulations, both compounds exhibit a single dominant
cluster (35: 88.7%; 37: 84.4%). The structure of 35 remains
mostly unchanged compared to its counterpart in DMSO and still
resembles a β-turn stabilized by the internal hydrogen bond
between Phe2 and Leu5 (Figure 8C, Table 2). On the other
hand, the helical shape of 37 appears slightly more pronounced
in water (Figure 9C, Table 3) than during the simulation in
DMSO.
In Figures SI 25 and SI 26 (see supporting information) we
Figure 8. Representative structures for 35 derived from clustering over the last 100 ns of distance-restrained MD in DMSO (A) and unrestrained simulations
in DMSO (B) and in water (C).
Figure 9. Representative structures for 37 derived from clustering over the last 100 ns of distance-restrained MD in DMSO (A) and unrestrained simulations
in DMSO (B) and TIP4Pew water (C).
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display the radial distribution functions (RDF) for chosen back-
bone atoms of 35 and 37, and Figure 10 highlights the most
relevant results. Such analysis reflects the solvation of specific
parts of the compounds and allows to further characterize their
behavior in water. In general, the plots show little to no structu-
ration of the solvent around the two molecules. Some hydrophilic
groups display clear narrow peaks at distances characteristic for
hydrogen bonds indicating stable interactions with water. How-
ever, the intensity of these peaks is rather small and the second
solvation shell is found further away. This shows that the global
solvation of the molecules is rather poor, which is in agreement
with the hydrophobic character of the side chains. All triazole
rings show similar solvation with N2 and N3 acting as hydrogen
bond acceptors. The RDFs corresponding to the hydrogen at-
oms of the rings (H4) show no particular structuration, empha-
sizing their low electrophilic character, as discussed by Marion
et al.[44] While most interactions for the two analyzed compounds
are matching, a clear difference appears for the solvation of two
backbone amino acid, i.e. Phe2 (O) and Leu5 (H). In 35, the
RDFs of both atoms (Figure 10) show that the first accessible
water molecules are found at distances greater than 4.0 Å.
compact conformation of 35 increases the hydrophobic charac-
ter of the molecule.
Conclusions
We have developed an efficient and robust synthesis route
leading to conformationally versatile peptidomimetic foldamers,
containing chiral 1,5-disubstituted triazoles in an alternating
fashion with amide bonds. Peptidomimetics with alternating
amide bonds and 1,5-disubstituted triazoles were obtained in a
building block approach: Six different homo- and heterochiral
building blocks Boc-Xaa[5Tz]Yaa-OBzl were synthesized in high
yields from chiral propargylamines and α-azido acids by Ruthe-
nium-catalyzed azide-alkyne cycloaddition (RuAAC) under mi-
crowave condition. Epimerization during the formation of the
peptide bonds could be prevented by a base-free carbodiimide
mediated preactivation.
The solid-state conformation of the homochiral tetramer Boc-
Val[5Tz]Ala-Leu[5Tz]Val-OBzl 34 and the structure from simu-
lated annealing in the gas phase without any constraint match
very well. The conformational preferences of the peptidotria-
zolamer hexamers Boc-Ala[5Tz]Phe-Val[5Tz]Ala-Leu[5Tz]Val-
OBzl 35 and Boc-D-Ala[5Tz]Phe-D-Val[5Tz]Ala-D-Leu[5Tz]Val-
OBzl 37 were elucidated by NMR measurements and MD simu-
lations in DMSO and in water. While the homochiral peptidotria-
zolamer 35 forms a compact β-turn-like structure, the heterochi-
ral peptidotriazolamer 37 with alternating chirality adopts a poly-
proline I-like conformation.
Acknowledgements
The authors gratefully acknowledge financial support from
Deutsche Forschungsgemeinschaft (DFG), Project SE 609/10-1,
Deutscher Akademischer Austauschdienst (DAAD), Project ID
57333947, the Leading National Research Centre (KNOW,
Poland), the BioNam: Bionanomaterials project, the Polish Min-
istry of Science and Higher Education (PMSHE) for the Faculty
of Chemistry of Wrocław University of Science and Technology,
Wroclaw Centre for Networking and Supercomputing
(http://wcss.pl), grant No. 197.
Figure 10. Radial distribution functions for chosen backbone atoms of 35 (top)
and 37 (bottom).
This is directly correlated with the fact that these atoms are
involved in formation of an intramolecular hydrogen bond, as
identified during the clustering analysis. In contrast, the extend-
ed conformation of 37 allows better solvation of the backbone in
comparison to 35. In consequence, the RDF of Phe2 (O) of 37
(Figure 10) has a clear narrow peak centered at 1.8 Å. The
backbone hydrogen atom Leu5 (H) in 37 is slightly more acces-
sible to water molecules, but does not show any particular struc-
turation, most likely due to large size of the aliphatic side chain
of this amino acid. Overall, this analysis shows that the confor-
mational differences between 35 and 37 highlighted during the
clustering analysis have an impact on their respective solvation
in water. While both compounds show quite poor solvation, the
extended conformation of 37 allows for interaction of all back-
bone hydrophilic groups with water. On the other hand, the
Conflict of Interest
The authors declare no conflict of interest.
Keywords: Peptidomimetics • Foldamers • Peptidotriazolamers
• RuAAC • Conformational Analysis
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Oliver Kracker, Jerzy Góra, Joanna
Krzciuk-Gula, Antoine Marion, Beate
Neumann, Hans-Georg Stammler, Anke
Nieß, Iris Antes, Rafał Latajka, and
Norbert Sewald*
((Insert TOC Graphic here; max. width: 11.5 cm; max. height: 2.5 cm))
Page No. – Page No.
Stereochemistry matters: While the conformation of the homochiral peptidotriazol-
amer Boc-Ala[5Tz]Phe-Val[5Tz]Ala-Leu[5Tz]Val-OBzl resembles that of a βVIa1
turn, the heterochiral peptidotriazolamer Boc-D-Ala[5Tz]Phe-D-Val[5Tz]Ala-D-
Leu[5Tz]Val-OBzl adopts a polyproline-like repetitive structure.
Title
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