Nucleoside derived amino acids (NDA) in foldamer chemistry: synthesis and conformational studies of homooligomers of modified AZT

Article abstract of DOI:10.1016/j.tetlet.2008.03.001

Homo short-oligomers of a novel trans-β-amino acid derived from AZT were synthesized and characterized. These adopt right-handed helical turns with their bases positioned systematically along the helix axis. These studies open up new possibilities for synthesizing nucleoside derived functional foldamers.

Full text of DOI:10.1016/j.tetlet.2008.03.001

Available online at www.sciencedirect.com
Tetrahedron Letters 49 (2008) 2969–2973
Nucleoside derived amino acids (NDA) in foldamer chemistry:
synthesis and conformational studies of homooligomers
of modified AZT
a,
a
b
*
S. Chandrasekhar , G. Pavan Kumar Reddy , M. Udaya Kiran ,
Ch. Nagesh a, B. Jagadeesh b,*
a Division of Organic Chemical Sciences, Indian Institute of Chemical Technology, Uppal Road, Tarnaka, Hyderabad 500 007, India
b Centre for Nuclear Magnetic Resonance, Indian Institute of Chemical Technology, Uppal Road, Tarnaka, Hyderabad 500 007, India
Received 4 December 2007; revised 28 February 2008; accepted 1 March 2008
Available online 6 March 2008
Abstract
Homo short-oligomers of a novel trans-b-amino acid derived from AZT were synthesized and characterized. These adopt
right-handed helical turns with their bases positioned systematically along the helix axis. These studies open up new possibilities for
synthesizing nucleoside derived functional foldamers.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords: Trans-b-amino acid; AZT; Oligonucleoside; NMR; Helical turn
Complex functions of proteins and peptides are strongly
associated with the orientation and relative positioning of
their functional groups along the structural backbone. In
the recent years, oligomers of unnatural peptidic
residues,1–3 particularly with b-amino acids4,5 and their
hybrids with natural L-amino acids (a) have emerged as
versatile structural templates (‘foldamers’) as these exhibit
predictable and well-defined secondary structures such as
helices, turns and strands, and offer a variety of possibilities
for orienting functional side-chains.
diversify our efforts from fused bicyclic sugar amino acids
to nucleoside derived amino acids (NDA), which are more
close in structure to the natural building blocks. Further-
more, oligomers of NDAs with specific conformational
preferences favour well-defined backbone folding and
geometrically positioned nucleobases, features that are
essential for improved molecular recognition.12,13 Chakra-
borty’s research group has reported amide linked (d-pep-
tide analogues) cyclic oligomers of thymidine-based
nucleoside amino acids.14 In their classic work, Gellman
et al. showed that the homooligomers of trans-b-ACPC
or heterooligomers with trans-APC residues adopt robust
left-handed 12-helix structures,15–19 stabilized by periodic
12-membered (NHi–COi-2) hydrogen bonding. Exploiting
both the predictable secondary structural features of b-pep-
tidic templates and the functional advantages of nucleo-
sides, herein we report the synthesis and folding
propensity of short homooligomers, trimer 9 and tetramer
10 of AZT derived trans-2,3-cyclic b-amino acid residue 2.
AZT 1, which is a drug of choice for the treatment of
AIDS, was chosen as the starting material. Through simple
We6–8 and other research groups9–11 have been engaged
in the synthesis and conformational studies of oligomers
based on cyclic beta sugar (furanose) amino acid building
blocks. The oligomers of these amino acids both in cis
and trans conformation have exhibited diverse helical pat-
terns in solution.6–9 These inspiring results prompted us to
*
Corresponding authors. Tel.: +91 40 2719 3210; fax: +91 40 2716 0512
(C.S.).
E-mail addresses: srivaric@iict.res.in (S. Chandrasekhar), bj@iict.
res.in (B. Jagadeesh).
0040-4039/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2008.03.001

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S. Chandrasekhar et al. / Tetrahedron Letters 49 (2008) 2969–2973
chemical transformations, 1 has furnished the trans-b
amino acid derivative 4, which possesses a bicyclic back-
bone but not the fused example as used in our earlier
approaches. A detailed conformational analysis was car-
ried out using circular dichroism (CD), NMR spectroscopy
and constrained molecular dynamics (MD) simulation
techniques, which have established that 9 and 10 adopt
robust helical folding in solution.
coupled under the same set of coupling conditions to
obtain smoothly tetramer 1022 in 40% yield. The purifica-
tion of this tetramer was cumbersome but could be
achieved by running a slow column on silica (Finer than
230–400 mesh). The white solid thus obtained has reason-
able shelf life. For our further studies, we have also synthe-
sized the shorter trimer 923 from dimer acid 7 and
monomer amine 3 using HOBt–EDCI (Scheme 2).21
Far-UV circular dicrosim (CD) spectroscopy data are
often used to derive the backbone folding pattern and
handedness of oligopeptides. The CD data for 200 lM
solutions of 9 and 10 (Fig. 1) in methanol exhibited distinct
secondary structural patterns with a minimum, zero-cross-
ing and maximum around 203, 212 and 224 nm, respec-
tively. Interestingly, the observed signatures are closely
comparable with those of 12-helical trans-b-ACPC oligo-
mers,15 but for the opposite handedness. The increased
ellipticity from trimer to tetramer is due to the increased
cooperative nature in higher oligomers in stabilizing the
helical structure, with the nucleation of the secondary fold-
ing being at a trimer level.
MeO
O
O
1
2
4
3
N
NH
O
O
BocHN
NDA
Initially, we synthesised the two functionalized dimers 7
and 8 to further couple to obtain tetramer 10. In this
endeavour, trimer 9 was obtained from coupling mono-
meric amine 3 with dimer acid 7. The commercially avail-
able AZT, 1 was oxidized in CH3CN/H2O (1:1) with
BAIB and TEMPO to furnish the azido acid20 which was
immediately esterified using CH2N2 to obtain the azido
methyl ester 2. This azido ester 2 was reduced via hydroge-
nation to produce the amino ester monomer 3 in 95% yield.
The second monomer 5 was obtained from 3 in two steps
involving the protection of the amino group as tert-butyl
carbamate 4 followed by mild ester hydrolysis using LiOH
in THF/H2O (3:1) (Scheme 1). The obtained monomers 3
and 5 were coupled using the well-established HOBt–EDCI
protocol21 to generate the protected dimer 6 in moderate
yield (60%). The required acid functionality in 6 was
released by treatment with LiOH to obtain dimer acid 7
in 88% yield.
Detailed NMR studies of 10 (1D, 2D-COSY, TOCSY,
ROESY) were carried out at 298 K in structure supporting
solvents, CD3OH and a mixture of CD3OH and CDCl3.
The backbone NH resonances were well dispersed (6.6–
8.6 ppm) in these solvents, indicative of secondary folding.
Whilst the structure was stable in methanol and 60:40
CD3OH/CDCl3 solvent mixture, complete assignment
was hampered due to overlap of the a and b proton reso-
nances of all the residues. However, they were relatively
better resolved in 10:90 CD3OH/CDCl3 (50 ll CD3OH
+550 ll CDCl3), in which the structure has been analyzed.
Even though these oligomers dissolve in DMSO-d6, we pre-
ferred to use methanol as solvent, since DMSO-d6 is likely
to disrupt/modify the native secondary structure. More-
over, methanol has been identified as a suitable structure
supporting solvent for trans-b-peptidic foldamers18 that
were investigated using CD and NMR spectroscopy. Pro-
tons in the bases were assigned from the intraresidual
NOEs and the sequential assignment of the residues was
achieved from the NHi–CHai-1 NOEs. Imide NHs in the
bases exhibited similar chemical shielding and all appeared
around 10.4 ppm as a broad peak. The complete chemical
shift assignment for 10 is given in Table 1. The temperature
coefficient (Dd/DT) studies of the NH proton resonances
carried out in this solvent suggested that 3NH (ꢀ5.5 ppb/
K) and 4NH (ꢀ7.0 ppb/K) are involved in moderate
hydrogen bonding while 1NH (ꢀ10.5 ppb/K) and 2NH
(ꢀ8.8 ppb/K) were solvent exposed. The marginally larger
temperature coefficients for 3NH and 4NH compared to
the usually observed values (>ꢀ4 ppb/K) are acceptable
for linear oligomers with intramolecular hydrogen bond-
ing.24 The relatively larger value observed for 4NH could
be due to a higher mobility of the less-constrained terminal
The second dimer 8 having a free amino functionality
was obtained from protected dimer 6 by treatment with
trifluoroacetic acid. The two dimers 7 and 8 were again
O
O
O
a, b
O
O
N
N
NH
NH
MeO
HO
O
c
O
N3
N3
2
1
O
O
O
O
e
O
O
N
N
NH
NH
HO
BocHN
MeO
O
O
R
5
3, R= NH2
4, R= NHBoc
d
Scheme 1. Reagents and conditions: (a) TEMPO, BAIB, CH3CN/H2O, rt,
12 h, 80%; (b) CH2N2, dry ether, 5 min, 10 °C, 90%; (c) Pd/C, MeOH, H2,
rt, 5 h, 95%; (d) (Boc)2O, Et3N, MeOH, rt, 10 h, 80%; (e) LiOH, THF/
H2O (3:1), 3 h, 0 °C, 95%.
3
residue. The large JNH–CbH values (ꢁ7.00 Hz, / ꢁ ꢀ115°)
for the trimer and tetramer show antiperiplanar arrange-
3
ment between these two protons and JCaH–CbH coupling

S. Chandrasekhar et al. / Tetrahedron Letters 49 (2008) 2969–2973
2971
O
O
O
O
O
O
O
O
R1HN
NH
OR
NH
NH
BocHN
OMe
a
a
5 + 3
7 + 3
O
O
3 days
60%
5 days
55%
T
T
T
T
T
6, R= Me, R1= Boc
7, R= H, R1= Boc
9 (Trimer)
b
c
8, R=Me, R1= H
O
T =
O
O
O
O
O
N
NH
NH
NH
NH
OMe
(Thymine)
BocHN
a
O
7 + 8
O
O
O
5 days
40%
T
T
T
T
10 (Tetramer)
Scheme 2. Reagents and conditions: (a) (i) EDCI, HOBt, DIPEA, DMF, rt; (b) LiOH, THF/H2O (3:1), 3 h, 0 °C, 88%; (c) TFA, CH2Cl2, 0 °C, 1 h.
of long range NOEs along the backbone turned out to be
ambiguous due to the overlapped resonances of the Ca
and Cb protons. However, in such cases, the NOEs involv-
ing the base methyl groups serve as suitable alternatives in
the structural elucidation of oligonucleotides and biomole-
cules.27,28 The observed long range inter-residue NOEs
between the methyl group protons of the thymidine bases
and the backbone or sugar ring protons, more specifically
1Me–4NH, 1Me–4H4, 1Me–4H1, 1Me–3H4, 2Me–3H2,
2Me–4H1 and 2Me–4H4 NOEs (Fig. 2b and a) indicate
that 10 adopts helical folding that favours 12-membered
NHi–COiꢀ3 hydrogen bonding.
Further proof for this possibility comes from the pres-
Fig. 1. CD spectra of trimer 9 and tetramer 10.
ence of a characteristic NOE between 3NH-Boc.19 The
(ꢂ5 Hz) corresponding to an N–Cb–Ca–CO angle, h25,26 of
adopted folding of the backbone closely positions the base
of the first residue (from the N-terminus) below the third
and fourth residues. It is worth noting that for tetramer
10, only two 12-membered hydrogen bonds involving
ꢂ+100°, which is consistent with that observed for 12-heli-
cal folds based on the other trans-b residues. The secondary
structure of tetramer 10 was established by a close analysis
3NH and 4NH each are possible. It is known that short
of the ROESY data, which showed convincingly that this
oligomers in principle exhibit various conformational pos-
homooligomer adopts right-handed helical folding, as
sibilities which can be in equilibrium and the NMR data
described below.
represent an average over these conformers. However, the
Whilst sufficient NOEs between adjacent residues could
present CD and NMR experimental data are suggestive
be identified to derive spatial connectivities, the assignment
Table 1
Chemical shift assignments for tetramer 10
Residue
1
NH
C1H(a)
C2H(b)
C3H(c)
C30H(c0)
C4H(d)
Hb
Meb
NHb
6.65 (d)
4.29 (d)
4.38 m
2.59 m
2.37 m
6.10 (t)
7.76 s
1.88 s
10.4 br s
JNH,H2 = 7.0
JH1,H2 = 5.0
JH4,H3 = 6.4
0
JH4,H3 = 5.0
2
3
4
8.61 (d)
JNH,H2 = 7.0
4.24 (d)
JH1,H2 = 5.1
4.61 m
4.65 m
2.55 m
2.49 m
2.27 m
2.48 m
2.49 m
2.40 m
6.20 (t)
8.06 s
8.10 s
8.05 s
1.90 s
1.96 s
1.96 s
10.4 br s
10.4 br s
10.4 br s
JH4,H3 = 6.2
JH4,H3 = 6.2
6.26 (t)
JH4,H3 = 6.2
JH4,H3 = 6.2
6.52 (dd)
8.47 (d)
JNH,H2 = 7.0
4.35 (d)
JH1,H2 = 5.0
8.64 (d)
4.65 (d)
4.70
JNH,H2 = 7.0
JH1,H2 = 2.3
JNH,H2 = 7.0
JH1,H2 = 2.3
JH4,H3 = 5.6
JH4,H3 = 8.7
Hb, Meb, and NHb indicate the protons present in the thymine base of the NDA.

2972
S. Chandrasekhar et al. / Tetrahedron Letters 49 (2008) 2969–2973
Fig. 2. (a) Schematic representation of the observed NOEs (in double headed black lines) and possible 12-membered hydrogen bonding (in single headed
colour curves) for tetramer 10. (b) Selected region of the ROESY spectrum of tetramer 10 supporting the helical fold. The cross-peaks are indicated with
the atomic groups involved. Example: 2Me–3H2 indicates the cross-peak between second residue base methyl to the third residue proton at position 2 (b).
of predominant population of 12-helical conformers in the
solvent medium chosen, which are also consistent with the
Gellman’s trans-b-peptidic foldamers.15–19
The intensities of the above mentioned inter and intra
residue NOEs were converted into distances by normaliz-
ing with respect to the NOE between the geminal protons
at the c position in each residue. These NOE derived dis-
tances (Table 2) and dihedral angles are used as constraints
in molecular dynamics simulations. The discover program
on Insight-II with CVFF force field has been used through-
out the simulations. Minimizations were determined first
with steepest decent, followed by conjugate gradient meth-
ods for a maximum of 10,000 iterations each or RMS devi-
ation of 0.001 kcal/mol, whichever was earlier. The energy
minimized structures were then subjected to MD simula-
tions using restraints, at 298 K. The molecules were ini-
tially equilibrated for 20 ps and subsequently went to a
1000 ps dynamics with 1 fs stepsize and trajectory sampling
for every 10 ps. A total of 100 sampled structures were gen-
erated and energy minimized. Figure 3 shows the superim-
posed structures of several such lower energy structures
obtained from MD simulations. The individual structures
showed about 2.5 residues per turn a geometrical feature
Table 2
Fig. 3. (a) Stereoview of superimposed minimum energy structures of
tetramer 10, (b) top view of the same along the helical axis showing the
orientation of the thymine bases with respect to the helix axis.
List of distance constraints used in MD simulations
˚
˚
NOEs
Distance (A) NOEs
Distance (A)
1
2
3
4
5
6
7
8
9
4NHM4H4
4NHM4H1
4NHM3H1
4HbM4H4
4HbM4H2
4H4M2Me
4H4M1Me
4H2M1Me
3NHM3H4
3NHM3H1
3NHM2H1
2.92–2.38
2.59–2.12
2.42–1.98
3.16–2.58
3.13–2.56
4.04–3.30
3.14–2.57
3.64–2.97
3.35–2.74
2.80–2.29
2.51–2.05
12 3HbM3H4
13 3HbM3H2
14 3H4 M1Me 3.28–2.68
2.87–2.35
2.87–2.35
that is relevant to 12-helical folding and an NH3–COBoc
-
˚
hydrogen bonding distance of ꢂ2.38 A. The bases are radi-
ally oriented along the helix axis (Fig. 3b) thereby provid-
ing enhanced accessibility of the biofunctional surface.
It is known that a specific helical folding is associated
with a cooperative nature of the constituent residues and
the structure attains better stability with the increase in
their population (greater than hexamer).7,29 Hence, it is
interesting and important to explore higher oligomers to
15 2NHM2H4
16 2NHM1H1
17 2HbM2H4
18 2HbM2H2
19 1NHM1H4
20 1NHM1H1
21 1HbM1H4
22 1HbM1H2
3.55–2.91
2.72–2.23
2.91–2.38
3.15–2.57
3.42–2.80
2.88–2.35
2.70–2.21
3.01–2.46
10
11

S. Chandrasekhar et al. / Tetrahedron Letters 49 (2008) 2969–2973
2973
9. Gruner, S. A. W.; Truffault, V.; Georg, V.; Locardi, E.; Stockle, M.;
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Mortensen, K.; Takahashi, M. J. Biol. Chem. 1998, 273, 15682.
13. Petrovykh, D. Y.; Dieste, V. P.; Opdahl, A.; Suda, H. K.; Sullivan, J.
M.; Tarlov, M. J.; Himpsel, F. J.; Whitman, L. J. J. Am. Chem. Soc.
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16. Wang, X.; Espinosa, J. F.; Gellman, S. H. J. Am. Chem. Soc. 2000,
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ascertain the observed conformational features. However,
the synthesis of higher oligomers beyond tetramers using
the conventional Boc-protected manual synthesis, was
hampered due to solubility problems. In order to derive
the details of specific helical folding in more populated
foldamers, solid phase synthetic methods have to be
employed, and work in this direction is in progress and will
be reported elsewhere.
In summary, we have discussed the synthesis and confor-
mational studies of AZT derived trans-b-amino acid short
trimer, and tetramer homooligomers, by using CD, NMR
and restrained MD techniques. The results have shown
the signatures of right-handed 12-helical turns that the
backbone adopts, which is consistent with Gellman’s
12-helices in the oligomers of trans-b-amino acids. In accor-
dance with the aim of our present work, the results encour-
age us to design functional oligonucleotides possessing well-
defined backbone folding and having their bases positioned
in a geometrically defined manner. However, detailed sec-
ondary structural features can only be obtained in hexamers
and beyond, and efforts towards this are in progress.30
19. Appella, D. H.; Christianson, L. A.; Klein, D. A.; Powell, D. R.;
Huang, X.; Barchi, J. J., Jr.; Gellman, S. H. Nature 1997, 387,
381.
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22. Spectroscopic data for compound 10: 1H NMR data listed in Table 1.
MS (ESI): m/z 1103 [M+Na]+.
Acknowledgement
23. Spectroscopic data for compound 9; 1H NMR (CDCl3, 500 MHz), d
G.P.K., thanks UGC, M.U.K. and C.N. thank CSIR,
New Delhi, India for research fellowships.
10.3 (br s, 3 NH), 8.57 (d, JNH,2 = 7.3 Hz, 1H), 8.50 (d, JNH,2
=
7.3 Hz, 1H), 8.06 (s, 2H), 7.91 (s, 1H), 6.63 (br s, 1H), 6.49 (t,
0
0
J4,3 = 8.4 Hz, J4;3 ¼ 5:6 Hz, 1H), 6.20 (t, J4,3 = 6.7 Hz, J4;3
¼
References and notes
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6:7 Hz, 1H), 6.12 (t, J4,3 = 6.5 Hz, J4;3 ¼ 6:5 Hz, 1H), 4.7 (m, 1H),
4.6 (m, 1H), 4.57 (d, J1,2 = 2.8 Hz, 1H), 4.34 (d, J1,2 = 4.2 Hz, 1H),
4.30 (m, 1H), 4.29 (d, J1,2 = 4.2 Hz, 1H), 3.81 (s, 3H), 2.26–2.54 (m,
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