Quantitative helix handedness bias through a single Hvs.CH3stereochemical differentiation

Article abstract of DOI:10.1039/d1cc01452h

A novel chiral aromatic δ-amino acid building block was shown to fully induce handedness in quinoline oligoamide foldamers with the possibility of further increasing the bias by combining multiples of these units in the same sequence. Through its incorporation within the helix, both N- and C-termini are still accessible for further functionalisation.

Full text of DOI:10.1039/d1cc01452h

ChemComm
View Article Online
View Journal
COMMUNICATION
Quantitative helix handedness bias through a
single H vs. CH3 stereochemical differentiation†
Cite this: DOI: 10.1039/d1cc01452h
Daniel Bindl,
Elisabeth Heinemann, Pradeep K. Mandal
and Ivan Huc
*
Received 17th March 2021,
Accepted 29th April 2021
DOI: 10.1039/d1cc01452h
rsc.li/chemcomm
A novel chiral aromatic d-amino acid building block was shown to Handedness bias is thus achieved through the stereochemical
fully induce handedness in quinoline oligoamide foldamers with the differentiation of a single main chain hydrogen atom and a
possibility of further increasing the bias by combining multiples of methyl group. This strong effect illustrates that the compactness
these units in the same sequence. Through its incorporation within of aromatic helices allows for stronger stereochemical differentia-
the helix, both N- and C-termini are still accessible for further tion than in peptidic helices in which a single alanine residue is
functionalisation.
unable to achieve complete handedness bias of an otherwise
achiral sequence.15
Handedness control in aromatic foldamer helices is of prime
We speculated that placing a stereogenic center within an
importance for their applications in circularly polarized aromatic helix may result in strong helix handedness bias due to
luminescence,1,2 enantioselective catalysis,3 and the diastereose- stereochemical constraints both above and below the chiral
lective recognition of chiral guests either within an internal group. Nevertheless, a few earlier attempts failed to reach quanti-
cavity4,5 or at their surfaces as, for example, proteins.6–10 Promot- tative handedness control, perhaps due to the fact that the chiral
ing minimal handedness bias is straightforward: most stereogenic groups were not themselves sufficiently helicogenic; that is, their
centers placed in the vicinity of the helix backbone will create an presence may cause a partial helicity disruption.16–19 Chiral
energy imbalance in favor of one or the other helix sense.11 Only monomer BRme was designed as a d-peptidic analogue of Q
1 kJ molÀ1 is required to elicit a 60/40 ratio. In contrast, bearing a stereogenic center at a position expected to be at the
quantitative handedness control, i.e. a diastereomeric excess of inner rim of the helix. Its achiral equivalent, 2-(2-aminophenoxy)-
at least 99%, has rarely been achieved because it requires a large acetic acid, has already been described,20 and homomeric
energy difference between P- and M-helical diastereomers. The sequences of this monomer have been shown to fold not into a
few examples reported concern aromatic oligoamide foldamers
derived from 8-amino-2-quinolinecarboxylic acid (Qn) (Fig. 1a)
possessing N- or C-terminal chiral moieties such as camphanyl
(Camph),12 oxazolylaniline (Oxaz)3,13 or b-pinene-derived pyridyl
(Pin)14 groups (Fig. 1b). All these moieties incorporate stereogenic
centers embedded within a cyclic system and form hydrogen
bonds with a main chain amide. Terminal functionalization by
a chiral moiety is often convenient, but it can hamper other
functionalizations, e.g. with a chromophore, a protein ligand, in
particular in the context of protein surface recognition.7–10 Here
we show that 2-(2-aminophenoxy)-propionic acid monomer BRme
(Fig. 2a) promotes quantitative handedness induction in water
when embedded within an aromatic oligoamide helix sequence.
Department of Pharmacy and Center for Integrated Protein Science,
¨
¨
Ludwig-Maximilians-Universitat, Butenandtstraße 5-13, Munchen 81377, Germany.
Fig. 1 (a) Chemical structures of Q and its oligomer Qn. Backbone
hydrogen bonds are indicated by red dashed lines. (b) N-terminal groups
(X) and C-terminal groups (Y), which are known to fully induce one
handedness in Qn oligomers.12–14
E-mail: ivan.huc@cup.lmu.de
† Electronic supplementary information (ESI) available: Synthetic and crystal-
lographic procedures. CCDC 2070816. For ESI and crystallographic data in CIF or
other electronic format see DOI: 10.1039/d1cc01452h
This journal is © The Royal Society of Chemistry 2021
Chem. Commun.

View Article Online
Communication
ChemComm
Fig. 2 (a) Foldamer building blocks used in this study. (b) Sequences that were synthesized to investigate the handedness induction properties of BRme
.
The chiral units (S)-Camph and BRme are shown in blue and red, respectively. (c–e) Crystal structure of oligomer 7 in side (c and d) and top (e) views
showing a canonical aromatic helix structure. Side chains and hydrogen atoms are omitted for clarity. The methyl group (red) and hydrogen atom (blue)
on the chiral center of BRme are represented as balls. The N- and C-terminus orientations are shown next to the respective structure.
canonical aromatic helix, but into
a herringbone-helical variety of B analogs bearing different chiral functionalities
structure.20,21 However, it is also known that Qn sequences can and/or side chains on the aromatic ring.
template the canonical helical folding of other monomers.19,22,23
Sequences 1–7 were designed and synthesized to study the
We thus endeavored to prepare an Fmoc protected version of BRme effect of one or two BRme units placed at various positions in
and to incorporate it into Qn oligomers to investigate its ability to sequences of variable lengths (Fig. 2b). Acidic QAsp monomers
bias helix handedness.
were introduced to provide solubility in aqueous media. To
Two new monomers, Fmoc-BRme-OH and Fmoc-BGly-OH, prevent too high solubility that could hamper crystal growth,
were synthesized from ortho-nitrophenol in 91% and 36% some QAla units were also included in the sequences so as to be
overall yield in four steps, respectively (Fig. S1, ESI†). The positioned on different faces of the helix. In some sequences,
stereogenic center of BRme was installed by condensing ethyl one QSem moiety containing a selenium atom was included to
(À)-L-lactate to 2-nitrophenol via a Mitsunobu reaction leading eventually enable the use of anomalous X-ray scattering,
to an inversion of the stereochemistry. A derivatization of the though this proved to be unneeded. Synthesis was performed
final Fmoc-BRme-OH with a chiral amine confirmed that on low loading Wang resin (100–200 mesh) using previously
the enantiomeric purity of the starting alcohol as given by the reported solid phase foldamer synthesis (SPFS) protocols.24,25
supplier (Z97.5%) was preserved throughout the whole synth- Fmoc acid building blocks were activated in situ by generating
esis (Section S2, ESI†). An unanticipated difficulty had to be the respective acid chlorides prior to coupling. Both Fmoc-
overcome during the preparation of Fmoc-BRme-OH and Fmoc-
B
B
Rme-OH and Fmoc-BGly-OH showed excellent coupling reactiv-
Gly-OH. The Mitsunobu product is a nitro ester that must ity. No noteworthy deletions occurred during SPFS. In a final
undergo nitro group reduction and ester saponification prior to step, the oligomers were cleaved from the resin and depro-
Fmoc installation. When performing reduction first, we found tected under acidic conditions. The crude oligomers were
that the amino-ester readily cyclizes into a six-membered purified using semi-preparative reverse phase HPLC with a
lactam. Even when saponification was carried out first, the basic ammonium acetate buffer as the mobile phase. As a
amino-acid was quantitatively converted into the same lactam result, all compounds were obtained as their respective ammo-
during nitro group hydrogenation (Fig. S1, ESI†). This unusual nium salts in good overall purified yields (8.4–51%; Section
reactivity might explain the scarce record of B oligomers in the S3.2, ESI†).
literature. Two different approaches were applied to circumvent
As outlined in the introduction, we expected that the strong
this side reaction. First, for BGly, the cyclisation was partially helicogenic nature of quinoline units would force the BRme and
prevented by using a bulky tert-butyl ester in the reduction step. BGly monomers into canonical helical folding despite their
For BRme, saponification was carried out first and a base steric demand and increased flexibility that results in a her-
(Na2CO3) was added during the hydrogenation to produce the ringbone helix for (BGly)n.20 NMR and circular dichroism (CD)
sodium carboxylate salt. This entirely prevented lactam for- spectroscopic data suggest that 1–7 all adopt a helical fold in
mation. These synthetic routes should be extendable to a water. They show the characteristic pattern of distinct amide
Chem. Commun.
This journal is © The Royal Society of Chemistry 2021

View Article Online
ChemComm
Communication
C-terminus of a sequence, as in 1 and 2. Since one helix turn
contains 2.5 units, the penultimate residue is entirely exposed
1
to the solvent on one of its faces. The H NMR spectrum of 1
shows one major set of signals (Fig. 3a), indicating a diaster-
eomeric ratio of (R)-M to (R)-P of at least 98/2, assuming that the
helix handedness inversion is slow on the NMR timescale, as
expected for a pentamer or any longer sequence.9 The negative
band observed by CD in the 300–450 nm region shows that the
M helicity is dominant.26 In the M helix, the asymmetric methyl
group of 1 should ‘stick out’ of the helix towards the solvent.
1
For 2, the H NMR spectrum shows two sets of signals with a
ratio of about 87/13 (Fig. 3a). The CD spectrum indicates that
the M helix is again dominant (Fig. 3b), which means that the
asymmetric methyl this time points towards the helix and not
to the solvent, since BRme is near the C terminus. The handed-
ness preference thus depends on the absolute stereochemistry
of BRme regardless of its position in the sequence, and not on
whether the methyl groups point towards the helix or the
solvent. We also note that the CD intensity does not correlate
well with the diastereomeric excess. The CD band at 380 nm
normalized per Q unit is more intense for 2 than for 1 although
the handedness bias is stronger for the latter (Fig. 3b). Indeed,
the CD intensity also depends on the number of consecutive Q
units and on the nature of substituents on each Q unit.
Compounds 3 and 4 contain one BRme unit flanked with
helix segments that span more than one turn. The asymmetric
center should thus have close contacts with aryl groups both
above and below. Sequence 4 was designed to be more flexible
than 3, due to additional achiral BGly units, including one
adjacent BRme. The higher flexibility arises from the reduced
Fig. 3 (a) Amide region of the 1H NMR spectra of sequences 1–7 in NH4OAc
buffer pH 8.5. For 2 and 5, major and minor sets of signals are marked with
red and blue circles, respectively. (b) CD spectra of compounds 1–7 in
NH4OAc buffer pH 8.5 between 300 and 500 nm. The molar extinction
(De) is normalized for the number of Q units for better comparability.
aromatic stacking surface and additional rotatable bonds in B
1
1
and aromatic signals in their H NMR spectra and an intense CD monomers as compared to Q. The H NMR spectra of 3 and 4
band in the 300–450 nm region that is typical for helically folded both show a single set of sharp signals (Fig. 3a). This indicates
quinoline oligomers with some handedness bias12,26 (Fig. 3 and quantitative handedness bias as far as NMR can detect. CD
Fig. S2, ESI†). These findings are corroborated by the strongly shows that 3 and 4 are M helical (Fig. 3b), as for 1 and 2.
downfield shifted 1H NMR signals (between À0.18 and À0.33 ppm) Quantitative handedness bias achieved by the stereochemical
of the CH3 protons of BRme units in the spectra of 3–7 (Fig. S2, differentiation between a hydrogen atom and a simple methyl
ESI†), as a result of the ring current effects of neighbouring Q units. group is remarkable and unprecedented. It probably results
For comparison, the signal of the same methyl group in from the very compact conformation of Qn helices that create a
Fmoc-BRme-OH is found at 1.56 ppm. Similar upfield shifts are large energy difference between the diastereomeric conformers.
observed for protons close to aromatic side chains in proteins.27
Furthermore, NMR and CD concur to show that the handed-
The helix conformation was also validated by the solid state ness bias for compound 3 is also quantitative in DMSO and
structure of 7, an analogue of 6 bearing an acetyl group instead MeOH (Fig. S4, ESI†).
of (S)-Camph (Fig. 2c–e). The structure shows canonical helical
Encouraged by these results, we challenged handedness bias
folding, with only left-handed helices present in the crystal. due to BRme through the introduction of an N-terminal cam-
This structure was solved despite the low resolution of the data phanyl group having an (S) configuration, that is, a configu-
(2.86 Å) by molecular replacement using an energy minimized ration antagonistic to that of BRme. In the absence of BRme, the
model (Section S4, ESI†). Molecular replacement is a common camphanyl group also biases handedness quantitatively and its
phasing method in protein crystallography where models are (S) configuration favors P helicity.12 Sequence 5 is an analog of
produced from related structures, but it has scarcely been used 3 where the terminal Ac has been replaced with (S)-Camph. Its
for smaller molecules.28 Successful molecular replacement in 1H NMR spectrum shows two sets of signals with a ratio of
the case of 7 is a highlight of the effectiveness of molecular 75/25 corresponding to the presence of two different diaster-
modelling at accurately predicting aromatic foldamer confor- eomeric conformations, (S,R)-M and (S,R)-P (Fig. 3a). The CD
mations, and represents an important methodological advance. spectrum shows a negative band in the range of 300–450 nm;
Helix handedness bias in solution was first assessed when this indicated that the major conformation has M handedness,
BRme was placed in the penultimate position to the N- or and thus that BRme imparts a stronger handedness induction
This journal is © The Royal Society of Chemistry 2021
Chem. Commun.

View Article Online
Communication
ChemComm
intensity observed for 5 in comparison to 3 reflects an 80/20 ratio of M-
to P-isomers
1 E. Merlet, K. Moreno, A. Tron, N. McClenaghan, B. Kauffmann,
Y. Ferrand and C. Olivier, Chem. Commun., 2019, 55, 9825–9828.
2 D. Zheng, L. Zheng, C. Yu, Y. Zhan, Y. Wang and H. Jiang, Org. Lett.,
2019, 21, 2555–2559.
than (S)-Camph (Fig. 3b). Because 3 and 5 contain the same
sequence of chromophores, their CD spectra should be directly
comparable. The relative CD intensities of 5 and 3 at their
maxima around 385 nm indicate an 80/20 ratio‡ of M- to
P-diastereomers for 5 (Fig. S3, ESI†), which matches the ratio
3 L. Zheng, D. Zheng, Y. Wang, C. Yu, K. Zhang and H. Jiang, Org.
Biomol. Chem., 2019, 17, 9573–9577.
1
observed by H NMR. Taking this ratio into consideration, an
4 G. Lautrette, B. Wicher, B. Kauffmann, Y. Ferrand and I. Huc, J. Am.
Chem. Soc., 2016, 138, 10314–10322.
5 S. Saha, B. Kauffmann, Y. Ferrand and I. Huc, Angew. Chem., Int. Ed.,
2018, 57, 13542–13546.
6 J. M. Alex, V. Corvaglia, X. Hu, S. Engilberge, I. Huc and
P. B. Crowley, Chem. Commun., 2019, 55, 11087–11090.
7 P. S. Reddy, B. Langlois d’Estaintot, T. Granier, C. D. Mackereth,
L. Fischer and I. Huc, Chem. – Eur. J., 2019, 25, 11042–11047.
8 M. Vallade, M. Jewginski, L. Fischer, J. Buratto, K. Bathany,
J.-M. Schmitter, M. Stupfel, F. Godde, C. D. Mackereth and I. Huc,
Bioconjugate Chem., 2019, 30, 54–62.
energy difference between the handedness bias induced by
BRme and Camph of about À3.4 kJ molÀ1 can be derived.
When two BRme units cooperate to bias handedness in the
same sequence, as in 6, we find that the effect of a terminal
(S)-Camph is completely reversed. A main species is observed
1
by H NMR, and CD confirms M handedness (Fig. 3). The CD
intensity of 6 also matches that of 7, which lacks the camphanyl
1
group. Minor signals in the H NMR spectra of 6 and 7 were
observed that can be assigned to the incomplete enantiomeric
purity of BRme arising from the enantiomeric purity of the
lactate precursor. A small amount of one or the other chiral B
unit may have (S) stereochemistry opposite to the (R) configu-
ration of BRme. In these cases, the effect of BRme and that of
BSme would cancel each other and the camphanyl group would
favor P helicity, leading to small amounts of (S,R,S)-P and
(S,S,R)-P diastereomeric conformers of 6 and (S,R)-P/M and
(R,S)-P/M conformers of 7, where the bias of Camph is missing.
In conclusion, quantitative handedness bias was achieved in
water, methanol, and DMSO by placing the new BRme monomer
within a quinoline helix. The energy difference was greater than
that generated by the Camph group. The bias could be further
enhanced by incorporating more than one BRme unit within the
same helix. The handedness bias was not complete only when
BRme was placed near the C-terminus. Full handedness control
could thus be achieved without any modifications at either the
N- or C-terminus, allowing for further functionalization at both
ends of the helix. Being able to avoid bulky handedness-
inducing groups at the N- or C-terminus will also be beneficial
to water solubility, as Camph, Pin, and Oxaz are all lipophilic.
These combined features will be useful for protein surface
recognition using helical foldamers. B units also provide a
new means to introduce side chains at the stereogenic center.
This prospect and the effect of multiple B units on the helix
geometry are being investigated and will be reported in due
course.
9 M. Vallade, P. Sai Reddy, L. Fischer and I. Huc, Eur. J. Org. Chem.,
2018, 5489–5498.
10 J. Buratto, C. Colombo, M. Stupfel, S. J. Dawson, C. Dolain,
B. Langlois d’Estaintot, L. Fischer, T. Granier, M. Laguerre,
B. Gallois and I. Huc, Angew. Chem., Int. Ed., 2014, 53, 883–887.
11 E. Yashima, N. Ousaka, D. Taura, K. Shimomura, T. Ikai and
K. Maeda, Chem. Rev., 2016, 116, 13752–13990; Z. Liu, X. Hu,
´
´
´
A. M. Abramyan, A. Meszaros, M. Csekei, A. Kotschy, I. Huc and
V. Pophristic, Chem. – Eur. J., 2017, 23, 3605–3615; A. Zhang,
Y. Han, K. Yamato, X. C. Zeng and B. Gong, Org. Lett., 2006, 8,
803–806.
12 A. M. Kendhale, L. Poniman, Z. Dong, K. Laxmi-Reddy,
B. Kauffmann, Y. Ferrand and I. Huc, J. Org. Chem., 2011, 76,
195–200.
13 Li Yang, M. Chunmiao, B. Kauffmann, L. Dongyao and Q. Gan, Org.
Biomol. Chem., 2020, 18, 6643–6650.
14 L. Zheng, Y. Zhan, C. Yu, F. Huang, Y. Wang and H. Jiang, Org. Lett.,
2017, 19, 1482–1485.
15 Y. Inai, Y. Kurokawa and N. Kojima, J. Chem. Soc., Perkin Trans. 2,
2002, 1850–1857; Y. Inai, Y. Kurokawa and T. Hirabayashi, Biopoly-
mers, 1999, 49, 551–564; M. D. Poli, M. D. Zotti, J. Raftery,
J. A. Aguilar, G. A. Morris and J. Clayden, J. Org. Chem., 2013, 78,
2248–2255; R. A. Brown, T. Marcelli, M. D. Poli, J. Sola and
J. Clayden, Angew. Chem., Int. Ed., 2012, 51, 1395–1399.
16 Z. Dong, G. P. A. Yap and J. M. Fox, J. Am. Chem. Soc., 2007, 129,
11850–11853.
17 H.-Y. Hu, J.-F. Xiang, Y. Yang and C.-F. Chen, Org. Lett., 2008, 10,
69–72.
18 E. Kolomiets, V. Berl and J.-M. Lehn, Chem. – Eur. J., 2007, 13,
5466–5479.
19 M. Kudo, V. Maurizot, B. Kauffmann, A. Tanatani and I. Huc, J. Am.
Chem. Soc., 2013, 135, 9628–9631.
20 M. Akazome, Y. Ishii, T. Nireki and K. Ogura, Tetrahedron Lett., 2008,
49, 4430–4433.
´
21 N. Delsuc, F. Godde, B. Kauffmann, J.-M. Leger and I. Huc, J. Am.
Chem. Soc., 2007, 129, 11348–11349.
22 C. Dolain, J. M. Leger, N. Delsuc, H. Gornitzka and I. Huc, Proc. Natl.
Acad. Sci. U. S. A., 2005, 102, 16146–16151.
Synchrotron data were collected at beamline P14 operated
by EMBL Hamburg at the PETRA III storage ring (DESY,
Hamburg, Germany). We thank Dr Saravanan Panneerselvam
for his assistance in using the beamline.
´
´
23 D. Sanchez-Garcıa, B. Kauffmann, T. Kawanami, H. Ihara,
M. Takafuji, M.-H. l. n. Delville and I. Huc, J. Am. Chem. Soc.,
2009, 131, 8642–8648.
24 B. Baptiste, C. Douat-Casassus, K. Laxmi-Reddy, F. Godde and
I. Huc, J. Org. Chem., 2010, 75, 7175–7185.
25 X. Hu, S. J. Dawson, Y. Nagaoka, A. Tanatani and I. Huc, J. Org.
Chem., 2016, 81, 1137–1150.
26 C. Dolain, H. Jiang, J.-M. Leger, P. Guionneau and I. Huc, J. Am.
Conflicts of interest
´
There are no conflicts to declare.
Chem. Soc., 2005, 127, 12943–12951.
27 C. C. Mcdonald and W. D. Phillips, J. Am. Chem. Soc., 1967, 89,
6332–6341.
28 G. W. Collie, K. Pulka-Ziach, C. M. Lombardo, J. Fremaux, F. Rosu,
M. Decossas, L. Mauran, O. Lambert, V. Gabelica, C. D. Mackereth
and G. Guichard, Nat. Chem., 2015, 7, 871–878.
Notes and references
‡The total CD intensity results from the sum of the CD signals of P- and
M-isomers, which have opposite signs. Therefore, the 40% lower CD
Chem. Commun.
This journal is © The Royal Society of Chemistry 2021