Orthanilic acid-promoted reverse turn formation in peptides

Article abstract of DOI:10.1039/c3cc40522b

Orthanilic acid (2-aminobenzenesulfonic acid, ^SAnt), an aromatic β-amino acid, has been shown to be highly useful in inducing a folded conformation in peptides. When incorporated into peptide sequences (Xaa- ^SAnt-Yaa), this rigid aromatic β-amino acid strongly imparts a reverse-turn conformation to the peptide backbone, featuring robust 11-membered-ring hydrogen-bonding.

Full text of DOI:10.1039/c3cc40522b

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Orthanilic acid-promoted reverse turn formation in
peptides†‡
Cite this: Chem. Commun., 2013,
49, 2222
Sangram S. Kale,a Gowri Priya,a Amol S. Kotmale,b Rupesh L. Gawade,c
Vedavati G. Puranik,c P. R. Rajamohananb and Gangadhar J. Sanjayan*a
Received 19th November 2012,
Accepted 30th January 2013
DOI: 10.1039/c3cc40522b
www.rsc.org/chemcomm
Orthanilic acid (2-aminobenzenesulfonic acid, SAnt), an aromatic
b-amino acid, has been shown to be highly useful in inducing a
folded conformation in peptides. When incorporated into peptide
sequences (Xaa-SAnt-Yaa), this rigid aromatic b-amino acid strongly
imparts a reverse-turn conformation to the peptide backbone,
featuring robust 11-membered-ring hydrogen-bonding.
b-Amino acids, the next higher homologues of a-amino acids,
provide considerable insight into designing synthetic peptides
with improved proteolytic and structural stability.1 Since
b-amino acids are stable against proteolytic degradation, they
are considered to be one of the key structural entities in drug
Fig. 1 Selected examples of b-amino acid building blocks.
design as well.1a The oligomers derived from b-amino acids attainable from some of these amino acid building blocks have
form stable secondary structures such as helices,2,3 sheets,4 found application in developing ligands for intervening protein–
etc., offering avenues for the structural exploration of diversely protein interactions.10 Among the various plausible intracatenary
functionalized novel b-amino acid building blocks and further hydrogen-bonded rings formed from b-amino acids, 11-membered
investigation of their conformational preferences.
hydrogen bonded ring is unusual and is considered as an
Recent years have witnessed the development of diversely alternative to Pauling’s a-helix (13-membered hydrogen bonded
functionalized b-amino acids with varying degrees of confor- ring)11a and also as the backbone extended analogue of the
mational freedom on the backbone, with respect to the parent typical b-turn (10-membered hydrogen bonded ring).11
b-alanine – the only naturally existing b-amino acid among the
Herein, we demonstrate orthanilic acid (2-aminobenzene-
S
b-amino acid family. Amongst the huge repertoire of b-amino sulfonic, Ant) as a unique aromatic b-amino acid with a very
acids, 2-aminocyclopropanecarboxylic acid (ACC),5 2-amino- high propensity to induce a b-turn-like conformation in
cyclopentanecarboxylic acid (ACPC),6 2-aminocyclohexane- peptides, displaying robust 11-membered-ring intramolecular
carboxylic acid (ACHC),7 and 2-aminobenzoic acid (anthranilic H-bonding interaction. With its amino and sulfonic acid
acid, Ant)8 (Fig. 1) attain special status owing to their backbone groups attached to the rigid aromatic framework, SAnt is
rigidity coupled with considerable scope for backbone function- characterized by well-defined torsional angles and constraints.
alization.9 It is noteworthy that the secondary structures It is noteworthy that sulfonic acid groups have a tetrahedral
geometry12 which is in stark contrast to the that of planar
a Division of Organic Chemistry, National Chemical Laboratory,
carboxyl groups usually found in amino acids. Thus, we
Dr Homi Bhabha Road, Pune 411 008, India. E-mail: gj.sanjayan@ncl.res.in;
envisioned that introduction of the rigid SAnt residue into
Fax: +91-020-2590-2629; Tel: +91-020-2590-2082
b Central NMR Facility, National Chemical Laboratory, Dr Homi Bhabha Road,
peptide sequences would not only cause rigidification of the
peptide backbone but also might have a dramatic outcome in
their conformation. Our efforts were also emboldened by the
general observation that sulfonamides as a class of compounds
are ushering into prominence owing to their potential use in
the development of drug candidates to treat different
ailments.13 Using several peptide sequences (Xaa-SAnt-Yaa),
we could unequivocally demonstrate the utility of SAnt in
Pune 411 008, India
c Center for Materials Characterization, National Chemical Laboratory,
Dr Homi Bhabha Road, Pune 411 008, India
† Dedicated to Prof. K. N. Ganesh on the occasion of his 60th birthday.
‡ Electronic supplementary information (ESI) available: General experimental
procedures, 1H, 13C, DEPT-135 NMR spectra and ESI mass spectra of all new
compounds. CCDC 910069–910074. For ESI and crystallographic data in CIF or
other electronic format see DOI: 10.1039/c3cc40522b
c
2222 Chem. Commun., 2013, 49, 2222--2224
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Fig. 2 Molecular structures (left) and their corresponding crystal structures (right) of peptides 1–6. Hydrogens, other than the ones involved hydrogen bonding, have
been omitted for clarity.
Table 1 Backbone torsional angles observed in crystals 1–6
strongly promoting the reverse-turn conformation featuring
11-membered robust intramolecular H-bonding, as clearly evident
Gly/Ala/Pro(i+1)
No. f (1) c (1)
SAnt(i+2)
from the solid and solution-state conformation of the peptides. The
robustness of the C-11 H-bonding has been tested by modulating
N- and C-termini of the peptides (1–6, Fig. 2). The N-terminus
modification has been achieved by altering the residues ranging
from the conformationally flexible glycine (Gly) to conformationally
restricted proline (Pro). Similarly, the C-terminus has also been
altered in an effort to understand the role of amino acid residues at
this position.
f (1)
y (1)
c (1)
o (1)
1
2
3
4
5
6
79.9(3)
86.7(2)
11.1(4)
9.0(2)
149.2(3) À5.0(3) À68.8(2) À93.5(3)
134.9(2) À5.9(2) À62.7(2) À90.7(1)
À75.7(4) À24.6(4) À102.5(4) À0.2(4)
58.0(3)
91.4(3)
À55.9(7) 139.8(4)
128.6(5) À4.8(6) À67.7(5) À71.2(4)
À85.6(2)
À6.8(3) À151.9(2)
5.4(3)
0.7(3)
69.9(2)
61.5(2)
63.9(2)
77.2(2)
—
—
—
All the designed peptides 1–5 commonly display two types of
intramolecular H-bonding: one type involving 6-membered
hydrogen bonding (C6 H-bonding) and another type involving
strong 11-membered hydrogen bonding (C11 H-bonding)
(Fig. 2). Whereas the C6 H-bonding is an intra-residual inter-
action, formed within SAnt itself, the C11 H-bonding is an
inter-residual strong H-bonding interaction, formed of
four residues between the CQO of the ith residue and the
sulfonamide NH of the (i + 3)rd residue, in the backward
direction (4 ’ 1), as seen in typical b-turns. The hydrogen-
bonding distances [d(HÁ Á ÁA)] of the C11 reverse turns in 1–5
were found to be ranging from 2.08 Å (1) up to 2.25 Å (5),
suggesting that the C11 H-bonded network is robust (Table 2).
Table 2 Hydrogen-bonding parameters observed in crystals 1–5a
Hydrogen bonding parameters (C11 H-bonding)
Compound no. HÁ Á ÁA (Å)
DÁ Á ÁA (Å)
D–HÁ Á ÁA (degree)
1
2
3
4
5
2.20
2.17
2.08
2.20
2.25
3.044(3)
2.992(2)
2.914(4)
2.887(6)
3.017(3)
169
160
162
137
148
a
Note: For further details, see ESI (S57–S70).
Finally, in order to understand the role of SAnt in folding, we
The hydrogen-bonding angle [D(D–HÁ Á ÁA)] varies from 1371 (4) made analog 6 which is devoid of the N-terminus arm essential
to 1691 (1). The dihedral angle descriptors of SAnt (Table 1), for the C-11 H-bonding. The crystal structure of 6 clearly reveals
with its twisted conformation, are informative. Of particular a folded conformation even in the absence of any inter-residual
note is the characteristic sulfonamide torsion (o) of SAnt H-bonding which clearly vindicates the pivotal role of SAnt with
observed in 1–5 which is in the range of 63.9(2)1 (5) to its twisted conformation in inducing folding (Fig. 2f). As in all
93.5(3)1 (1). It is noteworthy that in majority of the sulfonamide other cases, the usual intra-residual C-6 H-bonding of SAnt was
crystal structures reported in the literature,14 the sulfonamide observed in 6. The torsional angles and H-bonding parameters
torsion o has been found to be B901 (Æ30), which is in stark evidently provide insight into the conformational rigidity
S
contrast to the o of planar carboxamides (B1801 (Æ30)). The imparted by Ant, leading to the reverse turn conformation of
restriction of o to B901, obviously, imparts SAnt its twisted the peptides. It is noteworthy that 3 displays one more C11
conformation which is translated into the peptide backbone.
H-bonding between the CQO of the (i + 1) residue and NH of
c
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Fig. 3 Selected 2D nOe excerpts of 2 (a) and 5 (b) displaying C-11 membered hydrogen bonds (CDCl3, 400 MHz and 500 MHz, respectively).
the (i + 5) residue along with the regular reverse turn, described
SSK, GP and ASK thank CSIR, New Delhi, for research
earlier. This intriguing conformational feature singles out 3 fellowships. This work was funded by NCL-IGIB, New Delhi.
among others in the series, raising its prospects of being used
for the development of foldamers with perfect back-to-back
Notes and references
hydrogen bonding.2
The results of solution-state NMR studies firmly suggest that
the solid-state conformation is prevalent in the solution state as
well. The characteristic inter-residual nOe interactions supportive
of the folded conformation are the diagnostic long range inter-
residual dipolar couplings between the N- and C-termini groups.
Some of the selected nOes (Fig. 3a) that support the reverse-turn
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t
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t
Compounds 3 and 4 also display characteristic inter-residual nOes
supporting their folded conformation (ESI,‡ pages S42–S51). The
negligible chemical shift [Dd (NH) o 0.33 ppm (ESI,‡ pages
S29–S32)] observed in the DMSO-d6 titration experiments strongly
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(2-aminobenzenesulfonic acid, SAnt) is a strong reverse-turn
inducer when incorporated into peptide sequences (Xaa-SAnt-
Yaa). All the designed peptides containing SAnt displayed
11-membered-ring hydrogen bonding, formed in the backward
direction of the sequence as seen in native four-residue b-turns,
clearly vindicating the significance of the conformational rigidity
offered by SAnt to efficiently promote folding. The results of
diverse structural perturbations carried out in-and-around the
turn segment suggest that this reverse turn conformation is
robust. Comparison of the crystal structures15 clearly revealed
that it is the typical torsional constraint of orthanilic acid (SAnt)
that is primarily responsible for inducing folding in peptides.
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c
2224 Chem. Commun., 2013, 49, 2222--2224
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