Remarkable thermoresponsive nanofibers from γ-peptides

Article abstract of DOI:10.1039/c3cc45383a

Conformational analysis of γ-peptides composed of 4,4-gem-dimethyl γ-amino acids, their spontaneous self-assembly into nanofibrillar superstructures and remarkable thermoreversible gelation properties in various organic solvents are studied. This new gene

Full text of DOI:10.1039/c3cc45383a

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Remarkable thermoresponsive nanofibers
from c-peptides†
Cite this: Chem. Commun., 2013,
49, 9179
Sandip V. Jadhav and Hosahudya N. Gopi*
Received 16th July 2013,
Accepted 8th August 2013
DOI: 10.1039/c3cc45383a
www.rsc.org/chemcomm
Conformational analysis of c-peptides composed of 4,4-gem-dimethyl
c-amino acids, their spontaneous self-assembly into nanofibrillar super-
structures and remarkable thermoreversible gelation properties in
various organic solvents are studied. This new generation of c-peptides
may serve as potential templates to design advanced biomaterials.
Scheme 1 Synthesis of N-Boc-Aic from N-Boc-Aib. Reagents: (i) Ph3PQCHCOOEt,
THF; (ii) 10% Pd/C, MeOH; (iii) 1 N NaOH, MeOH.
Supramolecular self-assembly of peptides and proteins have
attracted considerable interest in recent years due to their utility in
the design and fabrication of nano-structures with tunable physical
and chemical properties. The low molecular weight gelators
(LMWGs), including short peptides, have been extensively studied
and have been shown to have applications in various fields includ-
ing tissue engineering, drug delivery, biomineralization, regenerative
medicine, wound healing and molecular electronics.1 Over the last
fifteen years b- and g-peptides have emerged as promising tools to
mimic the protein secondary structures.2 In addition, the self-
aggregation patterns of the peptides comprised of b- and g-residues
have also been recently investigated.3
Scheme 2 Schematic representation of Aic homooligomers and local torsional
variables of g-residues.
A literature survey reveals that g-peptides composed of g4-amino
acids, 3,3-dialkyl-g-amino acids and cyclic g-amino acids have a higher
tendency to fold into helical conformations.4 Recently, we demon-
strated the stable 12-helical conformations in a,g-hybrid peptides by
utilizing g4-residues.5 As a-amino isobutyric acid (Aib) has been
extensively utilized to design a-peptide helical secondary structures,6
we sought to investigate the structural properties of g-peptides
containing double homologated Aib, 4,4-gem-dimethyl substituted
g-amino acid (Aic, 4-amino isocaproic acid). The restriction in the f
and c conformational space of Aib forced the peptides to adopt stable
helical structures.6 Herein, we report the conformationally biased
unusual extended polar sheet type of structures from Aic homooligo-
mers (P1–P4), their unprecedented thermoreversible gels and supra-
molecular superstructures in various polar, apolar and protic solvents.
Fig. 1 (a) X-ray structures of Aic homooligomers P1 and P2; (b) assembly of P2
extended sheets into a double helical type of arrangement (view along a-axis)
and a polar extended sheet stabilized by intermolecular H-bonds (view along
b-axis). Arrow indicates direction of dipole.
Department of Chemistry, Indian Institute of Science Education and Research,
Dr. Homi Bhabha Road, Pune-411021, India. E-mail: hn.gopi@iiserpune.ac.in;
Fax: +91 (20) 2589 9790
† Electronic supplementary information (ESI) available: Experimental details,
NMR spectra, mass spectra and X-ray crystallography. CCDC 943867 and
943868. For ESI and crystallographic data in CIF or other electronic format see
DOI: 10.1039/c3cc45383a
In order to understand the conformational properties of
g4,4-peptides, the g4,4-amino acid, Aic, was synthesized starting
c
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from N-Boc-Aib aldehyde as shown in Scheme 1.5 All g4,4-peptides
(P1–P4, Scheme 2) were synthesized using standard DCC/HOBt
coupling conditions and purified by column chromatography.
Suitable X-ray quality single crystals of the peptides P1 and P2 were
obtained after the slow evaporation from the aqueous methanol
solution and their X-ray structures are shown in Fig. 1. Crystal
structure analysis reveals the presence of four molecules of P1 in
the asymmetric unit. Surprisingly, the peptides adopted parallel
extended planar structures stabilized by two intermolecular H-bonds.
Analysis reveals that one of the molecules in the asymmetric unit acts
as an H-bond donor and the other acts as an H-bond acceptor.
Further, the analysis of the single crystal structure of P2 suggested the
presence of two molecules in the asymmetric unit. In stark contrast to
the C9-/C14-helical conformations of the g-peptides with other g-amino
acids (g4-, g3,3-, etc.),4 the extended conformation of P2 observed in the
crystal structure is shown in Fig. 1. More intriguingly, the P2
molecules twisted around each other in an antiparallel fashion to
give a remarkable double helix type of structure (Fig. 1b) in the crystal
packing. Similar to P1, all NHs of P2 are involved in the H-bonding
with the carbonyls of the other peptide, leading to a polar sheet type
of character (Fig. 1b). This type of polar sheet character is generally
forbidden in the a-peptide b-sheets.7 The backbone conforma-
tions of P1 and P2 were measured by introducing two additional
torsional variables y1 (N–Cg–Cb–Ca) and y2 (Cg–Cb–Ca–C0) along with f
(C0–N–Cg–Cb) and c (Cb–Ca–C0–N) (Scheme 2). It is noteworthy that
torsional variables y1 and y2 mostly adopted gauche and anti con-
formations along N–Cg–Cb–Ca and Cg–Cb–Ca–C0, probably leading to
the formation of polar extended structures. In contrast, the g-residues
in the helical peptides adopted the g+, g+ conformations along
y1 and y2.2h,4,5 The torsional angles and the H-bond parameters
of P1 and P2 are given in the ESI.† Further, the 2D NMR
analysis of P2 in CDCl3 (6 mM) reveals the absence of long
Fig. 2 (A) NOEs observed in the 2D NMR are highlighted with double headed
arrows in both the peptides P3 and P4. No long distance NOEs were observed. (B)
Morphological studies of P3 and P4 xerogels. AFM images of P3 (a) and P4 (b)
xerogel in toluene. FE-SEM images of xerogel of P3 in 95% DMSO in water (c) and
toluene (d). FE-SEM images of P4 in MeOH (e) and toluene (f).
distance NH 2 NH or NH 2 CH NOEs and the strong inter- i + 1 residues, suggesting the extended type of planar structures
residue (i to i + 1) CaH 2 NH NOEs suggest the extended sheet similar to the tripeptide P2 (see ESI†). Further, the upfield shift of
type of conformation in solution similar to the g-peptides amide NHs with increasing temperature from ꢀ40 1C to +40 1C in
containing conformationally restricted cyclic g-residues.8
1H NMR experiments suggest their involvement in intermolecular
The unusual extended polar sheet conformation observed in P2 H-bonding (see ESI†). FT-IR studies were further undertaken to probe
encouraged us to grow the single crystals of tetra (P3) and penta (P4) the characteristic signature of intermolecular NHꢁꢁꢁOQC interactions
peptides in various solvents and solvent combinations. To our to understand the H-bond mediated aggregation in Aic oligomers.
surprise, both the peptides form a gel upon standing in methanol, The IR spectra of the xerogel of P3 and P4 are shown in the ESI.†
isopropanol, toluene and benzene. Astonished by the solvent gelation Peptides P3 and P4 displayed a sharp NH stretching vibration
of these peptides, we investigated the minimum concentration that is (nNH) at 3270 cmꢀ1 and 3273 cmꢀ1, respectively, supporting the
required for the gelation and found that 2 mg mLꢀ1 is sufficient to involvement of amide NHs in the intermolecular H-bonding.
get stable and transparent gels in toluene, benzene and isopropanol, Further, the amide I and amide II bands, which are directly related
and opaque gels in 95% DMSO in H2O, 95% DMF in H2O and 95% to the backbone conformations, were found to be 1637 cmꢀ1 and
diglyme in H2O. The inverted vial experiments depicting the gel 1555 cmꢀ1, respectively in P3. Similarly, P4 also displayed amide I
formation of peptide P3 in various solvents is shown in the ESI.† and amide II bands at 1640 cmꢀ1 and 1553 cmꢀ1, respectively. The
Similar results were also observed for P4 (see ESI†). All gels are found FT-IR values observed for the Aic homooligomers are in close agree-
to be thermoreversible: on heating all gels were transformed into ment with the values observed in the poly(Ala) infinite b-sheets.9 All
solution state and reconstituted upon cooling. As no gelation is these lines of experimental evidence support the extended confor-
observed in chloroform, we subjected these peptides to the 2D mations of Aic homooligomers.
NMR analysis in CDCl3 (3 mM) to understand their solution con-
Morphologies of the organogels of P3 and P4 were studied
formations. Unambiguous peak assignments were performed using using AFM (atomic force microscopy) and FE-SEM (field-emission
TOCSY, COSY and ROESY spectra. The NOEs observed in the ROESY scanning electron microscopy). The AFM images of P3 and P4
spectrum of P3 and P4 is shown in Fig. 2A. No NH 2 NH and long xerogels in toluene are shown in Fig. 2B–a and b, respectively. A
range NH 2 CH NOEs were observed in either of the peptides. network of entangled nanofibrillar aggregates of P3 and P4 is
However, strong CaH 2 NH NOEs were observed between i and observed in the AFM. The formation of ribbon-like one-dimensional
c
9180 Chem. Commun., 2013, 49, 9179--9181
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spontaneously self-assembled into nanofibers and displayed
unprecedented thermoreversible gelation in various solvents.
We are presently investigating the self-aggregation and confor-
mational properties of g-peptide heterooligomers with various
combinations of Aic and other g-residues. As short a-peptides
and low molecular weight gelator (LMWG)1 based biomaterials
have been finding applications in various fields, ranging from
tissue engineering to molecular electronics, the self-assembling,
gelation and self-healing properties of short Aic g-peptides may
offer a new generation of potential templates to design soft
biomaterials.
Fig. 3 Viscoelastic study: (A) strain sweep rheological study using 0.2% w/v of P3
in toluene at a constant oscillating frequency (10 rad sꢀ1) and temperature (20 1C);
(B) self-healing: temperature-ramp rheological study cycle using 0.2% w/v of P3 in
toluene with a ramp of 2 1C minꢀ1 at 0.05% strain and 10 rad sꢀ1 frequency.
We thank the Department of Science and Technology, Govt.
of India for financial support and NCL, Pune for the rheological
study. S. V. J. is grateful to CSIR, Govt. of India for a senior
fibrils in both the peptides P3 and P4, suggests a similar type of self- research fellowship.
aggregation. The FE-SEM micrographs of P3 xerogels in 95% DMSO
Notes and references
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