Artificial peptide-nanospheres self-assembled from three-way junctions of β-sheet-forming peptides

Article abstract of DOI:10.1021/ja052644i

Rational design of self-assembly of proteins, which plays pivotal roles in biology, is an important subject for biotechnology and also bottom-up nanotechnology. This paper has proposed a novel strategy for construction of artificial peptide-nanospheres by

Full text of DOI:10.1021/ja052644i

Published on Web 06/30/2005
Artificial Peptide-Nanospheres Self-Assembled from Three-Way Junctions of
â-Sheet-Forming Peptides
Kazunori Matsuura,* Kazuya Murasato, and Nobuo Kimizuka
Department of Chemistry and Biochemistry, Graduate School of Engineering, Kyushu UniVersity, Hakozaki 6-10-1,
Higashi-ku, Fukuoka 812-8581, Japan
Received April 23, 2005; E-mail: ma14tcm@mbox.nc.kyushu-u.ac.jp
Self-assembly of proteins such as formation of viruses, flagella,
microtubules, actin fibers, and amyloid fibrils plays pivotal roles
in biology.1 Rational design of such protein nano-assemblies will
give new applications not only in biotechnology but also in
nanotechnology.2 To date, fibrous peptide assemblies have been
designed based on the formation of â-sheets3 and leucine zippers.4
On the other hand, formation of spherical assemblies are limited
to the oil-/water-templated protein aggregates,5 self-assemblies of
amphiphilic peptides,6 aromatic dipeptides,7 and â-amyloid 1-40
peptides.8 These spherical assemblies are less exquisite compared
to the self-assembly of symmetric capsids in spherical viruses.1 For
example, icosahedral internal skeleton of tomato bushy stunt virus
is self-assembled from 60 C3-symmetric â-sheet subunits.9 Clathrin
lattice, which participates in receptor-mediated endocytosis, is a
polyhedral assembly of C3-symmetric triskelions formed in the
presence of Mg2+ 10
. The viral and clathrin’s self-assembly is thus
based on C3-symmetric architectures with directed intermolecular
interactions. This strategy would be generally applicable to the
design of artificial, nanosized polypeptide assemblies. We have
recently demonstrated self-assembly of DNA three-way junctions
with self-complementary sticky ends into mesoscopic spherical
DNA assemblies.11 In this paper, we extend the three-way
component design to polypeptides, and show a first example of
artificial C3-symmetric peptide conjugates that self-assemble into
viral-sized peptide nanospheres.
Figure 1. Synthesis of a C3-symmetric â-sheet peptide conjugate and
schematic illustration of the self-assembly.
As shown in Figure 1, the artificial C3-symmetric conjugate
Trigonal(FKFE)2 bears three â-sheet-forming peptides FKFEFKFE.3a
When these peptide arms form antiparallel â-sheets in water,
spontaneous self-assembly of peptide networks is expected. Trigo-
nal(FKFE)2 was prepared by coupling of thiol groups of CFK-
FEFKFE peptides with C3-symmetric iodoacetamidated core mol-
ecule 1 (Figure 1). The C3-symmetric 1 was synthesized by
condensation of trimesoyl chloride with N-Boc-ethylenediamine,
followed by deprotection and iodoacetylation. The 9-mer H-
CFKFEFKFE-OH peptide was prepared by standard solid-phase
synthesis using Fmoc chemistry (for FKFEFKFE). It was then
coupled with Boc-Cys(Trt)-OH and purified by reversed-phase
HPLC. The Trigonal(FKFE)2 conjugate was prepared by coupling
the 9-mer peptides with 1 in the presence of DIPEA (in degassed
DMF at -20 °C).12 The crude product was purified by reversed-
phase HPLC eluting with a linear gradient of acetonitrile/water (0/
100 to 60/40 over 60 min) containing 0.1% TFA (the isolated
yield: 8%). Isolation of the compound was confirmed by MALDI-
TOF-MS (m/z ) 4127 ([M + H]+)). The Trigonal(FKFE)2 was
soluble in acidic water but was hardly soluble in neutral and alkaline
water. Thus, the following experiments were conducted in acidic
aqueous dispersions (pH 3.3).
Figure 2. (A) CD spectrum of Trigonal(FKFE)2 (24 µM) in aqueous HCl
solution (pH 3.3) at 25 °C and (B) FT-IR transmission spectrum of
Trigonal(FKFE)2 cast from the acidic aqueous dispersion (24 µM, pH 3.3).
Substrate, BaF2.
Another negative peak at 200 nm is probably ascribed to an induced
CD for the benzene ring or amide bonds of the core molecule, since
such a CD peak was not observed for the component peptide
FKFEFKFE alone. FT-IR transmission spectrum of Trigonal-
(FKFE)2 cast on BaF2 plate from the acidic aqueous solution (24
µM, pH 3.3) showed a strong peak at 1630 cm-1 and a weak peak
at 1695 cm-1 (Figure 2B), which are characteristic of antiparallel
â-sheet structures.13 These results clearly indicate that Trigonal-
(FKFE)2 forms hydrogen-bond-mediated intermolecular assemblies,
since it is not possible for a single Trigonal(FKFE)2 molecule to
form â-sheets in an antiparallel manner.
The circular dichroism (CD) spectrum of the Trigonal(FKFE)2
(24 µM) in aqueous HCl (pH 3.3) showed negative peak at 219
nm, indicating the formation of â-sheet structures (Figure 2A).
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J. AM. CHEM. SOC. 2005, 127, 10148-10149
10.1021/ja052644i CCC: $30.25 © 2005 American Chemical Society

C O M M U N I C A T I O N S
In conclusion, supramolecular peptide-nanospheres are spontane-
ously self-assembled from artificial C3-symmetric â-sheet peptides.
The present C3-symmetric strategy would be widely applied to the
design of spherical bio-nanoassemblies. As the present C3-sym-
metric peptide can be chemically modified with desired functional
groups, we envisage their use in many fields such as gene carriers16
and as nanosized reactors.17
Acknowledgment. The present work is supported by a fund
from Sekisui Chemical Co., Ltd., a Grant-in-Aid for the 21st
Century COE Program from the Ministry of Education, Culture,
Sports, Science and Technology and by Grant-in-Aid for Scientific
Research (A) (16205028) from Japan Society for the Promotion of
Science.
Supporting Information Available: Experimental details, HPLC
chart, and MALDI-TOF-MS. This material is available free of charge
via the Internet at http://pubs.acs.org. This material is available free of
charge via the Internet at http://pubs.acs.org.
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Figure 3. AFM images (noncontact mode) of Trigonal(FKFE)2 (A) and
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