Regulation of β-sheet structures within amyloid-like β-sheet assemblage from tripeptide derivatives

Article abstract of DOI:10.1021/ja981363q

N-(1]-Trimethylammonioundecanoyl)-O,O'-didodecyl tripeptide bromides and N-(11-trimethylammonioundecanyl)-O-dodecyl tripeptide bromides formed a parallel β-Sheet structure when they aggregated in water and in CCl₄. The parallel β-sheet was dist

Full text of DOI:10.1021/ja981363q

12192
J. Am. Chem. Soc. 1998, 120, 12192-12199
Regulation of â-Sheet Structures within Amyloid-Like â-Sheet
Assemblage from Tripeptide Derivatives
Norihiro Yamada,*,† Katsuhiko Ariga,‡ Masanobu Naito,† Kazuhiro Matsubara,† and
Emiko Koyama†,§
Contribution from the Faculty of Education, Chiba UniVersity, 1-33 Yayoi-cho, Inage-ku, Chiba 263-8522,
Japan, and Graduate School of Materials Science, Nara Institute of Science and Technology (NAIST),
8916-5 Takayama, Ikoma, Nara 630-0101, Japan
ReceiVed April 20, 1998
Abstract: N-(11-Trimethylammonioundecanoyl)-O,O′-didodecyl tripeptide bromides and N-(11-trimethylam-
monioundecanyl)-O-dodecyl tripeptide bromides formed a parallel â-sheet structure when they aggregated in
water and in CCl4. The parallel â-sheet was distinguished from the antiparallel counterpart by Fourier transform
infrared spectroscopy because the former lacks a weak band at about 1690 cm-1 that is characteristic for the
latter. The FT-IR spectra of the aggregate in CCl4 remained unchanged if the solution was diluted to 0.01
mM, condensed to dryness, or heated to 60 °C, and hence, the â-sheet was easily formed and thermodynamically
stable. The parallel â-sheet was also possible to transform into an antiparallel â-sheet, for example, by mixing
with another tripeptide-containing amphiphile whose tripeptide part had an opposite direction. Transmission
electron microscope (TEM) and atomic force microscope (AFM) pictures revealed that the aggregate in CCl4
is a bundle of small filaments whose diameters are 70-80 Å. Developed interpeptide hydrogen bonding
should be formed along the long axis of the filament. The morphological structures and stable peptide
arrangements of the present assemblages are similar to those of the amyloid fiber whose accumulation causes
fatal diseases.
Inappropriate mutation of normal cellular proteins generates
pack according to a bilayer structure (Figure 1a and b) or an
interdigitated monolayer structure (Figure 1c) when they ag-
gregate.8 The former should produce a parallel â-sheet structure,
whereas the latter should form either a parallel or an antiparallel
â-sheet structure. On the other hand, an antiparallel â-sheet
would be predominantly formed if the two kinds of molecules
whose tripeptides had opposite directions were mixed with each
other. Thus, a highly ordered â-sheet structure which is the
characteristic property of the amyloid should be available from
the simple tripeptides.
Using Fourier transform infrared (FT-IR) spectroscopy, we
examined the detailed structure of the hydrogen bonding in the
assemblage of amphiphilic and hydrophobic tripeptide deriva-
tives. Morphological similarity between the assemblage and
the amyloid was also studied using a transmission electron
microscope (TEM) and an atomic force microscope (AFM).
fragmentary peptides or abnormal disease-causing isoforms
(prion peptides).1 The small peptides sometimes self-assemble
into an amyloid fibril, which accumulates in various internal
organs and causes functional diseases called amyloidosis.
Because the prion and the amyloid contain a rich amount of
â-sheet structure, it is important to constitute a â-sheet as-
semblage to understand the pathogenesis and the therapeutics
of these diseases. In this regard, we can find several model
studies on oligopeptides, for example, leucine-rich repeat
oligopeptides2 and ionic self-complementary oligopeptides.3
Aggeli suspects that all of the gel-forming oligopeptides could
also be applied to the amyloid model.4 However, it is not easy
to prepare the oligopeptide forming a desired secondary
structure, because all of the oligopeptides adopt an inherent
secondary structure depending on the kinds of component amino
acids and their number.
Experimental Section
In this paper, we report a new class of an amyloid model
using tripeptide derivatives shown in Chart 1. Because all of
the derivatives except for 8 formed an aggregate not only in
water but also in some nonpolar organic solvents,5-7 they should
Synthesis of Tripeptide Derivatives. N-tert-Butyloxycarbonyl-L-
amino acids (Boc-L-amino acids) were purchased from Calbiochem-
Novabiochem International, Inc., and 11-bromoundecanoic acid and
diethyl phosphorocyanidate (DEPC) from Wako Pure Chemical Ind.,
Ltd. All of the melting points were recorded on Yanaco micromelting
apparatus (MP-S3) and are uncorrected. The IR spectra were obtained
on a JASCO IRA-1 infrared spectrophotometer for the intermediates
and on a Nicolet 740 Fourier transform infrared (FT-IR) spectrophom-
eter for the final compounds according to the method described later.
† Chiba University.
‡ Nara Institute of Science and Technology.
§ Present Address: Department of Chemistry, University of Tsukuba,
Tsukuba, Ibaraki 305-8751, Japan.
(1) Prusiner, S. B. Science 1997, 278, 245-251.
(2) Symmons, M. F.; Buchanan, S. G. St. C.; Clarke, D. T.; Jones, G.;
Gay, N. J. FEBS Lett. 1997, 412, 397-403,.
1
Chemical shifts of the H NMR spectra were recorded on a Hitachi
(3) Zhang, S.; Rich, A. Proc. Natl. Acad. Sci. U.S.A. 1997, 94, 23-28.
(4) Aggeli, A.; Bell, M.; Boden, N.; Keen, J. N.; Knowles, P. F.; McLeish,
T. C. B.; Pitkeathly, M.; Radford, S. E. Nature 1997, 386, 259-262.
(5) Yamada, N.; Koyama, E.; Kaneko, M.; Seki, H.; Ohtsu, H.; Furuse,
T. Chem. Lett. 1995, 387-388.
R600 (60 MHz) or a JEOL GSX500 (500 MHz) spectrometer and are
given relative to tetramethylsilane (δ 0.00). All reactions were
(7) Yamada, N.; Koyama, E.; Maruyama, K. Kobunshi Ronbunshu 1995,
52, 629-638.
(8) Kunitake, T. Angew Chem., Int. Ed. Engl. 1992, 31, 709-726.
(6) Yamada, N.; Koyama, E.; Imai, T.; Matsubara, K.; Ishida, S. J. Chem.
Soc., Chem. Commun. 1996, 2297-2298.
10.1021/ja981363q CCC: $15.00 © 1998 American Chemical Society
Published on Web 11/11/1998

Amyloid-Like â-Sheet Assemblage
J. Am. Chem. Soc., Vol. 120, No. 47, 1998 12193
Chart 1
spot); IR (CCl4 past) ν(N-H) 3380, 3280 cm-1, νas(CH2) 2920 cm-1
νs(CH2) 2840 cm-1, ν(CdO, ester) 1740 cm-1, ν(CdO, urethane) 1710
cm-1, ν(CdO, amide I) 1680 cm-1, δ(N-H, amide II) 1530 cm-1
ν(C-C-O, ester) 1180 cm-1
,
,
.
To the solution of Boc-Ala-Glu(OC12H25)2 (2.90 g, 4.43 mmol) in
10 mL of dichloromethane was added 28.7 g of 25% HBr-acetic acid
at once under stirring with ice cooling. After gas evolution ended (4
hs), the mixture was neutralized with saturated sodium hydrogen
carbonate with ice cooling. The mixture was poured into chloroform,
washed three times with saturated sodium hydrogen carbonate and with
saltwater, dried over dry magnesium sulfate, and evaporated to dryness
in vacuo. The residual oil was solidified in a freezer and was
recrystallized from petroleum ether to give a colorless powder (H-Ala-
Glu(OC12H25)2, 1.90 g, 77%): mp 41.0-45.0 °C; TLC Rf 0.30 (one
spot); IR (CCl4 past) ν(N-H) 3600-3200 cm-1 (broad), νas(CH2) 2940
cm-1, νs(CH2) 2840 cm-1, ν(CdO, ester) 1750 cm-1, ν(CdO, amide
I) 1680 cm-1, δ(N-H, amide II) 1540 cm-1. The bands attributed to
the urethane linkage completely disappeared.
Figure 1. Different possibilities of arranging tripeptide-containing
molecules. (a) Aqueous bilayer structure with a single component. (b)
Reversed bilayer structure with a single component in organic media.
(c) Interdigitated monolayer structure with a single component either
in water or in organic media.
Into a mixture of H-Ala-Glu(OC12H25)2 (1.80 g, 3.24 mmol) and
Boc-L-Ala-OH (0.66 g, 3.49 mmol) in 50 mL of dry THF was added
Et3N (0.35 g, 3.50 mmol) at once. Et3N‚HCl immediately precipitated.
To the mixture was added a solution of diethyl phosphorocyanidate
(0.57 g, 3.50 mmol) in 30 mL of dry THF within 15 min with stirring
at a temperature below 5 °C. The mixture was allowed to stand
overnight at room temperature and was filtered and evaporated to
dryness in vacuo. The residual oil was dissolved in chloroform, washed
three times with saturated sodium hydrogen carbonate, washed with
saltwater, and evaporated. The residue (Boc-Ala-Ala-Glu(OC12H25)2)
was not solidified and was used without further purification, because
the TLC result (Rf 0.41, one spot) indicated the presence of no other
monitored by thin-layer chromatography (Merck TLC aluminum sheet
silica gel 60 GF254; developer, 10:1 chloroform-methanol). Elemental
analyses (C, H, and N) were performed at the Analysis Center, Chiba
University.
The double-chain, tripeptide-containing amphiphiles 1-7 were
prepared by a stepwise condensation of Boc-L-amino acid to didodecyl-
L-glutamate. The following method was used for the synthesis of 2
and 8 (an intermediate of 2) which is the general synthetic procedure.
N-[N-[N-(11-Bromoundecanoyl)-L-alanyl]-L-alanyl]-O,O′-didodec-
yl-L-glutamate (8). Into a mixture of didodecyl-L-glutamate hydro-
chloride9 (3.00 g, 5.77 mmol) and Boc-L-Ala-OH (1.14 g, 6.00 mmol)
in 50 mL of dry THF was added Et3N (0.61 g, 6.03 mmol) at once.
Et3N‚HCl immediately precipitated. To the mixture was added a
solution of diethyl phosphorocyanidate (0.98 g, 6.01 mmol) in 30 mL
of dry THF within 15 min with stirring at a temperature below 5 °C.
The mixture was allowed to stand overnight at room temperature and
was filtered and evaporated to dryness in vacuo. The residual oil was
dissolved in chloroform, washed three times with saturated sodium
hydrogen carbonate and with saltwater, evaporated, and recrytallized
from a methanol-water mixture to give a colorless powder (Boc-Ala-
Glu(OC12H25)2, 3.57 g, 95%): mp 42.0-45.0 °C; TLC Rf 0.40 (one
components: IR (neat) ν(N-H) 3440, 3340 cm-1, νas(CH2) 2940 cm-1
νs(CH2) 2860 cm-1, ν(CdO, ester) 1740 cm-1, ν(CdO, urethane) 1720
cm-1, ν(CdO, amide I) 1680 cm-1, δ(N-H, amide II) 1530 cm-1
ν(C-C-O, ester) 1180 cm-1
,
,
.
Into a mixture of the Boc-Ala-Ala-Glu(OC12H25)2 (2.35 g, 3.23
mmol) in 10 mL of dichloromethane was added 15.3 g of 25% HBr-
acetic acid was added at once under stirring with ice cooling. After
gas evolution ended (3 hs), the mixture was neutralized with saturated
sodium hydrogen carbonate with ice cooling. The mixture was poured
into chloroform, washed three times with saturated sodium hydrogen
carbonate and with saltwater, dried over dry magnesium sulfate, and
evaporated to dryness in vacuo. A colorless oil was obtained (H-Ala-
(9) Kunitake, T.; Nakashima, N.; Hayashida, S.; Yonemori. K. Chem.
Lett. 1979, 1413-1416.

12194 J. Am. Chem. Soc., Vol. 120, No. 47, 1998
Yamada et al.
Ala-Glu(OC12H25)2, 1.87 g, 93%): TLC Rf 0.43; IR (neat) ν(N-H)
3260 cm-1, νas(CH2) 2920 cm-1, νs(CH2) 2840 cm-1, ν(CdO, ester)
54H, 27CH2 in long chain), 1.72(brs, 2H, N+CH2CH2), 1.95 (brs, 2H,
OCOCH2CH2 in Glu), 2.05-2.50 (brm, 4H, NCOCH2 and OCOCH2),
3.20-3.65 (brs, 11H, CH2N+(CH3)3), 3.75-4.20 (brm, 8H, 2COOCH2
and 2CH2 in Gly), 4.20-4.75 (brm, 1H, CH in Glu). Anal. Calcd for
C47H91N4O7Br‚3/2H2O: C, 60.62; H, 10.17; N, 6.02. Found: C, 60.59;
H, 10.15; N, 5.77.
1740 cm-1, ν(CdO, amide I) 1640 cm-1, δ(N-H, amide II) 1540 cm-1
.
The IR spectra contained no Boc-Ala-Ala-Glu(OC12H25)2, and TLC
indicated that no other impurity was included; hence, the material was
used in the next synthesis without further purification.
Into a mixture of H-Ala-Ala-Glu(OC12H25)2 (1.87 g, 2.99 mmol) and
11-bromoundecanoic acid (0.95 g, 3.95 mmol) in 30 mL of dry THF
was added Et3N (0.36 g, 3.59 mmol) at once. To the mixture was
added a solution of diethyl phosphorocyanidate (0.58 g, 3.59 mmol)
in 30 mL of dry THF within 15 min with stirring at a temperature
below 5 °C. The mixture was allowed to stand overnight at room
temperature, filtered, dried over dry magnesium sulfate, and evaporated
to dryness in vacuo. The residual solids were dissolved in chloroform,
washed with saltwater, evaporated, and recrytallized twice from
methanol to give a colorless powder 8 (1.66 g, 64%): mp 85.5-87.5
3: recrytallized from acetone containing a small amount of ethanol;
a colorless powder; mp 191.0-193.5 °C; IR (1 mM of CCl4 solution)
ν(N-H) 3280 cm-1, νas(CH2) 2927 cm-1, νs(CH2) 2853 cm-1, ν(Cd
O, ester, see Figure 4) 1734 cm-1, ν(CdO, amide I, see Figure 4) 1632
cm-1, δ(N-H, amide II, see Figure 4) 1546 cm-1; 1H NMR (60 MHz,
CDCl3) δ 0.75-1.05 (br, 18H, 2CH3 in long chain and 4CH3 in Val),
1.05-1.85 (brs, 56H, 28CH2 in long chain), 2.00 (brs, 2H, OCOCH2CH2
in Glu), 1.85-2.45 (brm, 6H, NCOCH2, OCOCH2 and 2CH(CH3)2 in
Val), 3.30-3.60 (brs, 11H, CH2N+(CH3)3), 3.80-4.20 (brm, 4H,
2COOCH2), 4.20-4.70 (brm, 3H, 3CH in Glu and Val). Anal. Calcd
for C53H103N4O7Br‚2H2O: C, 62.64; H, 10.58; N, 5.40. Found: C,
62.40; H, 10.43; N, 5.30.
°C; TLC Rf 0.66; IR (10 mM of CCl4 solution) ν(N-H) 3281 cm-1
,
νas(CH2) 2928 cm-1, νs(CH2) 2853 cm-1, ν(CdO, ester, see Figure 2)
1738 cm-1, ν(CdO, amide I, see Figure 2) 1630 and 1688 cm-1, δ(N-
4: recrytallized from acetone; a colorless powder; mp 180.0-184.0
1
H, amide II, see Figure 2) 1527 cm-1; H NMR (60 MHz, CDCl3) δ
°C; IR (1 mM of CCl4 solution) ν(N-H) 3281 cm-1, νas(CH2) 2926
cm-1, νs(CH2) 2854 cm-1, ν(CdO, ester, see Figure 4) 1736 cm-1
,
0.90 (brt, 6H, 2CH3 in long chain) 1.05-1.80 (brm, 60H, 27CH2 in
long chain and 2CH3 in Ala), 1.87 (brs, CH2CH2Br), 2.00-2.60 (brm,
6H, NCOCH2 and OCOCH2CH2 in Glu), 3.41 (t, J ) 7 Hz, 2H, CH2Br),
3.90-4.30 (m, 4H, COOCH2), 4.30-4.90 (brm, 3H, CH in Ala), 6.43
(d, J ) 7 Hz, 1H, NH in Ala), 6.85-7.40 (brm, 2H, NH in Ala and in
Glu). Anal. Calcd for C46H86N3O7Br: C, 63.28; H, 9.93; N, 4.81.
Found: C, 63.33; H, 10.31; N, 4.89.
ν(CdO, amide I, see Figure 4) 1632 cm-1, δ(N-H, amide II, see Figure
4) 1546 cm-1; 1H NMR (60 MHz, CDCl3) δ 0.60-1.05 (br, 18H, 2CH3
in long chain and 2CH(CH3)CH2CH3 in Ile) 1.10-1.90 (brs, 64H,
29CH2 in long chain, 2CH(CH3)CH2CH3 in Ile and OCOCH2CH2 in
Glu), 2.00-2.60 (br, 6H, OCOCH2, NCOCH2 and 2CH(CH3)CH2CH3
in Ile), 3.20-3.60 (brs, 11H, CH2N+(CH3)3), 3.80-4.40 (brm, 4H,
2COOCH2), 4.40-4.70 (brm, 3H, 3CH in Glu and Ile). Anal. Calcd
for C55H107N4O7Br‚1H2O: C, 63.86; H, 10.62; N, 5.42. Found: C,
63.83; H, 10.56; N, 5.47.
N-[N-[N-(11-Trimethylammonioundecanoyl)-L-alanyl]-L-alanyl]-
O,O′-didodecyl-L-glutamate Bromide (2). The resultant bromide 8
(1.34 g, 1.87 mmol) was dissolved in 100 mL of dry THF. Dry Me3N
gas was bubbled into the solution under vigorous stirring with ice
cooling until the total volume of the solution slightly increased. The
flask was stoppered and allowed to stand for 1 week at room
temperature in the dark. The solvent and excess Me3N were removed
under reduced pressure. In this procedure, simultaneous use of an amine
trap and a hood was required because Me3N has an extremely bad odor.
The residual solid was twice recrystallized from acetone to give a
colorless powder (2, 1.24 g, 71%): mp 103.5-106.0 °C; TLC Rf 0
(there is no developed component); IR (1 mM of CCl4 solution) ν(N-
H) 3399 cm-1, νas(CH2) 2921 cm-1, νs(CH2) 2852 cm-1, ν(CdO, ester,
see Figure 2) 1747 and 1728 cm-1, ν(CdO, amide I, see Figure 2)
1647 and 1631 cm-1, δ(N-H, amide II, see Figure 2) 1560 and 1534
cm-1; 1H NMR (500 MHz, CDCl3) δ 0.82 (t, 6H, 2CH3 in long chain),
1.15-1.40 (brm, 54H, 24CH2 in long chain and 2CH3 in Ala), 1.48-
1.60 (m, 6H, 2COOCH2CH2 and NCOCH2CH2), 1.77 (brs, 2H, N+-
CH2CH2), 1.90-2.10 (m, OCOCH2CH2 in Glu), 2.20 (brs, 2H,
NCOCH2), 2.25-2.40 (m, 2H, OCOCH2 in Glu), 3.37 (s, 9H,
N+(CH3)3), 3.60 (brs, 2H, N+CH2), 3.92-4.05 (m, 4H, 2COOCH2),
4.31-4.37 (m, 1H, CH in Glu), 4.42 (brs, 2H, CH in Ala), 7.44 (brs,
1H, NH in Ala), 7.59 (brs, 1H, NH in Ala), 7.83 (brs, 1H, NH in Glu).
Anal. Calcd for C49H95N4O7Br‚1H2O: C, 61.94; H, 10.29; N, 5.90.
Found: C, 61.92; H, 10.08; N, 5.87.
5: recrytallized from acetone; a colorless powder; mp 109.0-111.5
°C; IR (1 mM of CCl4 solution) ν(N-H) 3282 cm-1, νas(CH2) 2925
cm-1, νs(CH2) 2854 cm-1, ν(CdO, ester) 1737 cm-1, ν(CdO, amide
I) 1637 cm-1, δ(N-H, amide II) 1542 cm-1 1H NMR (500 MHz,
;
CDCl3) δ 0.88 (t, J ) 7 Hz, 6H, 2CH3 in long chain), 1.10-1.40 (brm,
48H, 24CH2 in long chain), 1.51-1.53 (m, 6H, 2COOCH2CH2 and
NCOCH2CH2), 1.77 (brs, 2H, N+CH2CH2), 2.00-2.25 (m, 2H,
OCOCH2CH2 in Glu), 2.35-2.50 (m, 4H, NCOCH2 and OCOCH2 in
Glu), 3.00-3.20 (m, 4H, CH2Ar in Phe), 3.28 (s, 9H, N+(CH3)3), 3.50-
3.58 (brm, 2H, N+CH2), 4.00-4.10 (m, 4H, 2COOCH2), 4.45-4.51
(m, 1H, CH in Glu), 4.64-4.80 (m, 2H, 2CH in Phe), 7.15-7.25 (m,
10H, Ar-H in Phe), 7.50 (brs, 1H, NH in Glu or Phe), 8.35 (brs, 1H,
NH in Glu or Phe), 8.65 (brs, 1H, NH in Glu or Phe). This spectrogram
was shown in ref 5. Anal. Calcd for C61H103N4O7Br‚2H2O: C, 65.39;
H, 9.63; N, 5.00. Found: C, 65.31; H, 9.66; N, 4.99.
6: recrytallized from acetone; a colorless powder; mp 100.0-101.8
°C; IR (1 mM of CCl4 solution) ν(N-H) 3280 cm-1, νas(CH2) 2926
cm-1, νs(CH2) 2854 cm-1, ν(CdO, ester) 1737 cm-1, ν(CdO, amide
I) 1632 cm-1, δ(N-H, amide II) 1543 cm-1; 1H NMR (60 MHz, CDCl3)
δ 0.88 (t, J ) 7 Hz, 6H, 2CH3 in long chain), 1.10-1.50 (brm, 54H,
27CH2 in long chain), 1.77 (brm, 2H, N+CH2CH2), 2.00-2.80 (brm,
10H, OCOCH2CH2 in Glu, SCH2CH2 in Met and NCOCH2), 2.10 (s,
6H, 2SCH3), 3.40 (s, 9H, N+(CH3)3), 3.50 (s, 2H, N+CH2), 3.90-4.27
(brm, 4H, 2COOCH2), 4.38-4.90 (brm, 3H, 3CH in Glu and Met),
7.17 (brs, 1H, NH in Glu or Met), 7.57 (brs, 1H, NH in Glu or Met),
7.75 (d, J ) 6 Hz, 1H, NH in Glu or Met). Anal. Calcd for
C53H103N4O7S2Br‚H2O: C, 59.47; H, 9.89; N, 5.23. Found: C, 59.22;
H, 10.06; N, 5.10.
Other Amphiphiles. Other double-chain, tripeptide-containing
amphiphiles 1, 3-7 were prepared in the same manner as that described
above. The single-chain, tripeptide-containing amphiphiles 9-11 were
synthesized using the corresponding dodecyl amino acid instead of
didodecyl-L-glutamate, which procedure has been reported elsewhere.7
The amphiphile 12 whose tripeptide part had an opposite direction was
synthesized as follows. 10-Bromodecyloxy-L-phenylalaninate hydro-
chloride was condensed with Boc-Phe-Phe-OH in the same manner as
that described above. After the protection group was removed, the
resulting oil was allowed to react with tridecanoic acid, which was
followed by quaternization with Me3N as described above. The
analytical data for these final compounds are shown below.
7: recrytallized from acetone; a colorless powder; mp 145.0-152.0
°C; IR (1 mM of CCl4 solution) ν(N-H) 3264 cm-1, νas(CH2) 2927
cm-1, νs(CH2) 2855 cm-1, ν(CdO, ester, see Figure 2) 1738 cm-1
,
ν(CdO, amide I, see Figure 2) 1634 cm-1, δ(N-H, amide II, see Figure
1
2) 1543 cm-1; H NMR (60 MHz, CDCl3) δ 0.70-1.10 (brm, 18H,
2CH3 in long chain, 2CH3 in Val and 2CH3 in Leu), 1.15-1.60 (brm,
54H, 27CH2 in long chain), 1.60-2.10 (br, 6H, N+CH2CH2, CH2CH-
(CH3)2 in Leu, OCOCH2CH2 in Glu), 2.10-2.70 (brm, 4H, NCOCH2
and OCOCH2 in Glu), 2.75-3.15 (brm, 2H, CH(CH3)3 in Val and
CH2CH(CH3)2 in Leu), 3.38 (s, 9H, N+(CH3)3), 3.40 (s, 9H, N+(CH3)3),
3.50 (s, 2H, N+CH2), 3.90-4.30 (brm, 4H, 2COOCH2), 4.30-4.70 (m,
3H, 3CH in Glu, Val, and Leu), 6.65 (brs, 1H, NH in Glu, Val, or
1: recrystallized from acetone; a colorless powder; mp 88.0-91.0
°C; IR (1 mM of CCl4 solution) ν(N-H) 3366 and 3319 cm-1, νas(CH2)
2924 cm-1, νs(CH2) 2852 cm-1, ν(CdO, ester, see Figure 2) 1727 and
1709 cm-1, ν(CdO, amide I, see Figure 2) 1677, 1663 and 1649 cm-1
,
1
δ(N-H, amide II, see Figure 2) 1537 and 1513 cm-1; H NMR (60
MHz, CDCl3) δ 0.80 (brt, 6H, 2CH3 in long chain), 1.00-1.65 (brm,

Amyloid-Like â-Sheet Assemblage
J. Am. Chem. Soc., Vol. 120, No. 47, 1998 12195
Leu), 6.79 (brs, 1H, NH in Glu, Val, or Leu), 7.10 (brs, 1H, NH in
Glu, Val, or Leu). Anal. Calcd for C54H105N4O7Br‚3H2O: C, 61.40;
H, 10.59; N, 5.30. Found: C, 61.61; H, 10.65; N, 5.32.
TEM Measurements. The staining of aggregates in organic solvents
was carried out in the same manner as that described in ref 10. A
sample solution (2 mM) was dropped onto a carbon-coated TEM grid.
Excess liquid was blotted off and the residue dried in vacuo. The
observation was carried out with a JEOL JEM 100S transmission
electron microscope.
AFM Measurements. The AFM observation was carried out for
the cast films from different solutions on freshly cleaved mica. The
AFM images were taken by a Nanoscope IIIa (Digital Instruments,
Santa Barbara, CA) in a tapping mode in air. According to the
manufacturer, the normal spring constant was 20-100 N/m. Drive
frequency was around 300 kHz. Scanning speed was at a line frequency
of 1 Hz with 256 pixels per line.
9: recrytallized from a mixture of chloroform and hexane; a colorless
powder; mp 159.2-164.9 °C; IR (CCl4 solution) ν(N-H), ν(C-H),
ν(CdO, ester) 1740, ν(CdO, amide I), δ(N-H, amide II), ν(C-C-
1
O, ester) cm-1; H NMR (60 MHz, CDCl3) δ 0.80 (brt, 3H, CH3 in
long chain), 1.00-1.95 (brm, 42H, 18CH2 in long chain and 2CH3 in
Ala), 2.00-2.40 (brm, 2H, NCOCH2), 3.35 (s, 9H, N+(CH3)3), 3.55
(brs, 2H, N+CH2), 3.90-4.20 (brm, 2H, COOCH2), 4.20-4.70 (brm,
3H, 3CH in Ala), 7.35 (brs, 1H, NH in Ala or Glu), 7.53 (brs, 1H, NH
in Ala or Glu), 7.67 (brs, 1H, NH in Ala or Glu). Anal. Calcd for
C35H69N4O5Br‚9/4H2O: C, 56.32; H, 9.93; N, 7.51. Found: C, 56.37;
H, 9.65; N, 7.76.
Results and Discussion
10: recrytallized from acetone; a colorless powder; mp 178.3-182.7
°C; IR (1 mM of CCl4 solution) ν(N-H) 3279 cm-1, νas(CH3) 2961
The amphiphiles used in the study formed an aggregate not
only in water but also in some nonpolar organic solvents.5-7
On the other hand, they formed no aggregate in polar solvents
such as ethanol and chloroform. The aggregation behavior can
be monitored by vibrational spectroscopy because the aggrega-
tion of the peptides is accompanied by band shifts from a
nonbonding to a characteristic amide bonding mode.6 The
nonbonding amide modes at 3412-3426 cm-1 (amide A),
1659-1678 cm-1 (amide I), and 1503-1516 cm-1 (amide II)
were observed in the CHCl3 solutions without respect to the
tripeptide structure, whereas the amide bonding modes were
observed in aqueous solutions and in CCl4 solutions (Table 1).
The FT-IR spectra of these bonding bands were divided into
four types as shown in Figure 2. The type I spectrum is the
most ordinary structure in the present study, where amide A
absorbed at 3271-3285 cm-1, amide I only absorbed at 1631-
1638 cm-1 and amide II at 1536-1547 cm-1. Amide A and
amide I are more than 100 and 20 cm-1 lower, respectively,
than those observed in CHCl3 solution. Amide II, a deformation
mode of N-H, is 20 cm-1 higher than that observed in CHCl3.
Furthermore, the half bandwidths of the bonding amide I bands
were ca. 12 cm-1, which were 30 cm-1 smaller than that of the
nonbonding amide I bands in CHCl3 solution (ca. 50 cm-1).
These results show that a strong and orderly hydrogen bond
was formed in the aggregate of these amphiphiles. According
to a previous report, N-deblocked-heptavaline, heptaisoleucine,
and heptaphenylalanine, which adopted a parallel â-sheet
alignment in the crystal, showed the same amide mode because
amide I and amide II appeared at 1635-1639 cm-1 and 1540-
1549 cm-1, respectively, and no other peaks appeared in the
region.11 Therefore, the tripeptide-containing amphiphiles
except for 1 and 2 should form the parallel â-sheet structure in
the aggregate without respect to whether the solvent is aqueous
or organic.
cm-1, νas(CH2) 2927 cm-1, νs(CH3) 2874 cm-1, νs(CH3) 2855 cm-1
,
ν(CdO, ester) 1737 cm-1, ν(CdO, very small intensity of nonbonding
amide I) 1661 cm-1, ν(CdO, bonding amide I) 1632 cm-1, δ(N-H,
1
amide II) 1545 cm-1; H NMR (60 MHz, CDCl3) δ 0.60-1.05 (brm,
21H, CH3 in long chain and 3CH(CH3)CH2CH3 in Ile), 1.03-2.00 (brm,
42H, 18CH2 in long chain and 3CH(CH3)CH2CH3 in Ile), 2.10-2.45
(brs, 2H, NCOCH2 and OCOCH2), 3.05 (brs, 3H, 3CH(CH3)CH2CH3
in Ile), 3.40 (s, 11H, CH2N+(CH3)3), 3.80-4.24 (m, 2H, COOCH2),
4.24-4.75 (brm, 3H, 3CH in Ile), 7.00-7.80 (br 3H, NH in Ile). Anal.
Calcd for C44H87N4O5Br‚5/4H2O: C, 61.84; H, 10.56; N, 6.56. Found:
C, 61.92; H, 10.79; N, 6.44.
11: recrytallized from ethyl acetate; a colorless powder; mp 97.0-
101.0 °C; IR (1 mM of CCl4 solution) ν(N-H) 3282 cm-1, νas(CH2)
2926 cm-1, νs(CH2) 2854 cm-1, ν(CdO, ester) 1738 cm-1, ν(CdO,
amide I) 1638 cm-1, δ(N-H, amide II) 1544 cm-1; 1H NMR (60 MHz,
CDCl3) δ 0.85 (brt, 3H, CH3 in long chain), 1.05-1.70 (brm, 34H,
17CH2 in long chain), 1.81 (brs, 2H, N+CH2CH2), 2.00-2.50 (brm,
2H, NCOCH2), 2.80-3.20 (brm, 6H, 3CH2Ar), 3.30 (s, 9H, N+(CH3)3),
3.40-3.70 (brm, 2H, N+CH2), 3.75-4.20 (brm, 2H, COOCH2), 4.25-
4.80 (brm, 3H, 3CH in Phe), 6.55 (brs, 1H, NH in Phe), 6.90- 7.30
(m, 16H, 3Ar-H in Phe and NH in Phe), 7.75 (brs, 1H, NH in Phe).
Anal. Calcd for C53H81N4O5Br‚5/2H2O: C, 65.01; H, 8.85; N, 5.72.
Found: C, 65.03; H, 8.94; N, 5.75.
12: recrytallized from ethyl acetate; a colorless powder; mp 90.5-
93.2 °C; IR (1 mM of CCl4 solution) ν(N-H) 3271 cm-1, νas(CH2)
2926 cm-1, νs(CH2) 2854 cm-1, ν(CdO, ester, see Figure 6) 1739 cm-1
,
ν(CdO, very small intensity of nonbonding amide I, see Figure 6) 1663
cm-1, ν(CdO, bonding amide I, see Figure 6) 1638 cm-1, δ(N-H,
1
amide II, see Figure 6) 1541 cm-1; H NMR (500 MHz, CDCl3) δ
0.88 (t, J ) 7 Hz, 3H, 2CH3 long chain) 1.15-1.33 (m, 26H, 13CH2
in long chain), 1.36 (brs, 2H, N+CH2CH2CH2), 1.42-1.50 (m, 2H,
NCOCH2CH2), 1.51-1.58 (brm, 2H, COOCH2CH2), 1.72 (brs, 2H,
N+CH2CH2), 2.12 (t, J ) 7.3 Hz, 2H, NCOCH2), 2.96 (d, J ) 7 Hz,
2H, CH2Ar), 2.99 (d, J ) 7.3 Hz, 2H, CH2Ar), 3.03-3.13 (m, 2H,
CH2Ar), 3.30 (s, 9H, N+(CH3)3), 3.42-3.48 (m, 2H, N+CH2), 3.95-
4.15 (m, 2H, COOCH2), 4.56 (d, t, J ) 7 Hz, J ) 7 Hz, 1H, CH in
Phe), 4.57 (d, t, J ) 7 Hz, J ) 7 Hz, 1H, CH in Phe) 4.69 (d, t, J )
7 Hz, J ) 7 Hz, 1H, CH in Phe), 6.67 (d, J ) 8 Hz, 1H, NH in Phe),
7.01 (d, J ) 7.4 Hz, 1H, NH in Phe), 7.10-7.30 (m, 15H, 3Ar-H in
Phe), 7.39 (d, J ) 8.2 Hz, 1H, NH in Phe). Anal. Calcd for
C53H81N4O5Br‚1/2H2O: C, 67.49; H, 8.76; N, 5.94. Found: C, 67.27;
H, 8.95; N, 5.88.
The formation of the parallel â-sheet is explained from a
morphological aspect. Possessing a hydrophilic head and
hydrophobic chains, the amphiphiles formed a normal bilayer
structure (Figure 1a) in water.8 An interdigitated monolayer
structure (Figure 1c) is unfavorable in water because the
hydrophilic ammonium head is buried in the hydrophobic chain
region. The structure is possible only in the organic media.
However, we observed a fibrous structure in CCl4 solutions of
the tripeptide-containing amphiphiles by transmission electron
microscopy. For example, Figure 3 is the TEM image of 2 in
three different organic solvents. Small fibers observed in these
images have a 70-80 Å diameter. Because the expanded length
of the molecule is 45 Å, the diameter corresponds to the
FT-IR Measurements. The sample solutions were prepared by
sonication. In cases of binary systems, components were mixed in
CHCl3, and the solvent was evaporated. The mixed film thus obtained
was dissolved in CCl4 by sonication. The sample solution was
sandwiched between the CaF2 windows with a 19-µm spacer for CHCl3
and CCl4 solutions and a 50-µm spacer for the aqueous solution. The
CaF2 windows were mounted in a temperature-controlled flow-through
cell (Harrick Scientific Co., TFC-M19). The IR spectra were recorded
on a Nicolet 740 Fourier transform infrared spectrometer with a TGS
detector at 25 ( 0.5 °C. Two hundred interferograms were co-added
and Fourier transformed with one level of zero filling to yield spectra
(10) Ishikawa, Y.; Kuwahara, H.; Kunitake, T. J. Am. Chem. Soc. 1994,
116, 5579-5591.
(11) Toniolo, C.; Palumbo, M. Biopolymers 1977, 16, 219-224.
with a high S/N ratio at a resolution of 4 cm-1
.

12196 J. Am. Chem. Soc., Vol. 120, No. 47, 1998
Yamada et al.
Table 1. Absorption Frequencies of Tripeptide Molecules in Different Media
frequencies/cm-1
amide I
tripeptide molecules
solventsa
CCl4
water
CHCl3 (5 mM)
CCl4
water
CCl4
CCl4
CHCl3 (16 mM)
CCl4
spectral typeb
amide A
amide II
1
IV
E
N
III
III
I
I
N
I
3366, 3319
c
3414
3299
c
3280
3281
3412
3282
c
1677, 1663, 1649
1650
1661
1647, 1631
1647, 1632
1632
1632
1668
1637
1638
1537, 1513
1549
1503
1560, 1534
1561, 1534
1546
1546
1518
1542
1538
2
3
4
5
water
I
6
7
CCl4
CHCl3
CCl4
I
N
I
3280
3420
3285
c
1631
1664
1634
1635
1543
1516
1543
1544
water
I
8
CHCl3 (20 mM)
CCl4 (10 mM)
water
CCl4
water
N
II
3420
3277
insoluble
precipitate
c
1678
1504
1527
insoluble
precipitate
1547
1688, 1630
insoluble
precipitate
1634
9
I
10
CHCl3
CCl4
water
N
I
I
3426
3279
c
1659
1632
1633
1505
1545
1548
11
12
CCl4
water
CCl4
I
I
I
3282
c
3271
1636
1637
1638
1545
1536
1541
a Concentrations of the sample solutions were 1 mM for CHCl3 and CCl4 solution and 20 mM for aqueous solutions. b N and E denote a nonbonding
amide mode and a specific mode except for a â-sheet structure (see text), respectively, and the others (I-IV) are classified according to Figure 2.
c It is impossible to subtract the solvent absorption in this region because of the strong ν(O-H) band of water. Measurement temperature was 25.0
( 0.5 °C. The other conditions are cited in parentheses.
The type II spectrum is attributed to a typical antiparallel
â-sheet structure where amide I absorbs at 1630 cm-1, together
with a weak band at 1688 cm-1, and amide II at 1527 cm-1
.
The major band of amide I in the antiparallel â-sheet absorbed
at a slightly lower frequency in comparison with that in the
parallel â-sheet structure. The essential difference between the
structures is whether the weak band at about 1690 cm-1 is
present. This weak band has been inferred from the results of
a theoretical calculation for the antiparallel â-sheet polypep-
tides11,12 and observed in crystalline heptaalanine, heptanorva-
line, heptaleucine, heptamethionine, and hepta-S-methylcysteine
containing an antiparallel â-sheet structure.11 In cases of the
heptapeptides, the major amide I band also appeared at a lower
frequency (1630-1633 cm-1) than that of the peptides forming
a parallel â-sheet structure (1635-1639 cm-1). The type II
spectrum was only observed in the CCl4 solution of the
hydrophobic Ala-Ala-Glu derivative 8. In this case, the
hydrophobic 8 should pack according to an interdigitated
structure (Figure 1c).
The type III spectra were characteristic of Ala-Ala-Glu-
containing amphiphile 2 in water and CCl4, where the bands
attributed to the ester, amide I and amide II split into two peaks.
For example, the amide I band split into 1647 and 1631 cm-1
(Figure 2). The presence of the major peak at 1631 cm-1 and
the absence of the weak band at about 1690 cm-1 indicate the
formation of a parallel â-sheet conformation. However, the
band split, particularly at the ester carbonyl, suggests that one
of the amide groups forms a hydrogen bond with either an R-
or a γ-ester group. We call this specific amide mode a
pseudoparallel â-sheet structure.
Figure 2. FT-IR spectra of four different types of secondary structures
of tripeptide derivatives: type I (parallel â-sheet), CCl4 solution of 7
(1 mM); type II (antiparallel â-sheet), CCl4 solution of 8 (10 mM);
type III (pseudoparallel â-sheet), CCl4 solution of 2 (1 mM); type IV,
CCl4 solution of 1 (1 mM). All spectra were measured at 25.0 ( 0.5
°C.
thickness of a reversed bilayer structure (Figure 1b) in which
the components are slightly tilted. This result means that the
local structure of the aggregate in organic solvents did not adopt
an interdigitated structure (Figure 1c) but a reversed bilayer
structure. The normal and reversed bilayer structures contain
the same molecular alignment in their half-layer and hence
contained the same parallel â-sheet structure.
The type IV spectrum was observed in the CCl4 solutions of
Gly-Gly-Glu-containing amphiphile 1. The multiple peaks
assigned to the amide I band meant that the amphiphile did not
form any â-sheet structures. The amphiphile gave different
(12) Bandekar, J. Biochim. Biophys. Acta 1992, 1120, 123-143.

Amyloid-Like â-Sheet Assemblage
J. Am. Chem. Soc., Vol. 120, No. 47, 1998 12197
Figure 4. Temperature dependence of FT-IR spectra of 3 in CCl4
solution.
Figure 5. AFM image of the air-dried cast film from CCl4 solution of
2.
similar to that of the amyloid filaments. On the other hand,
isotropic CHCl3 solution, which showed a nonbonding amide
mode, gave the type I spectra when the solvent was removed
to dryness (Table 2). In particular, that of the amphiphile 2
was characteristic. The CHCl3 solution of 2 produced the type
I spectrum, a parallel â-sheet, after air-drying even if the air-
dried film from CCl4 produced the type III spectrum, a
pseudoparallel â-sheet. Therefore, we obtained a pseudoparallel,
a parallel, and an antiparallel â-sheet, by modifying the chemical
structure, changing the solvent, and/or drying the solutions.
The light intensity at the nonbonding amide I band (1659-
1678 cm-1) within the spectra of the â-sheets (Figures 2 and
4) shows that the â-sheets should extend for a long period. In
fact, transmission electron microscope (TEM) pictures revealed
well-developed filaments whose diameters are 70-80 Å (Figure
3). The direction of all of the tripeptide chains should be
oriented perpendicularly to the axis of the filament, which
corresponds with the Alzheimer’s â-peptide in which the peptide
is perpendicular to the fiber axis.13 These filaments assembled
into a bundle, which can be observed three-dimensionally using
AFM. Although AFM is now widely used for various objects
on a µm to sub-µm scale as well as for observations at the
molecular level,14 observations at the sub-µm level have been
Figure 3. TEM images of the assemblage of 2 in three different
solvents (scale bar; 1000 Å). (a) CCl4, (b) benzene, (c) cyclohexane.
spectra depending upon the conditions. In this aggregate, the
H-bonding could be formed two-dimensionally because glyci-
nate residues possessed no sterically hindered side group.
The respective FT-IR spectra (types I, II, and III) of the
solutions of the tripeptide-containing amphiphiles remained
unchanged when the solutions were diluted or condensed to
dryness. For example, the type I amide mode of Phe-Phe-Glu
5 was unchanged in CCl4 solution at 25 °C even if the
concentration decreased to 0.01 mM which is the lower limit
of detection. Therefore, the critical concentration of the
aggregate formation of 5 is lower than 0.01 mM in CCl4 at 25
°C. Heating also caused no spectral change. Figure 4 is an
example of the temperature dependence of FT-IR spectra
observed in the CCl4 solution of 3. The spectra remained
unchanged if the temperature was raised to 60 °C, although we
did not measure the sample at a still higher temperature because
of the difficulty with temperature control. These results show
that the aggregate possesses a thermal stability, which is very
(13) Kirschner, D. A.; Inouye, H.; Duffy, L. K.; Sinclair, A.; Lind, M.;
Selkoe, D. J. Proc. Natl. Acad. Sci. U.S.A. 1987, 84, 6953-6957.
(14) Shao, Z.; Mou, J.; Czajkowsky, D. M.; Yuan, J.-Y. AdV. Phys. 1996,
45, 1-86.

12198 J. Am. Chem. Soc., Vol. 120, No. 47, 1998
Yamada et al.
Table 2. Absorption Frequencies of Air-Dried Cast Films of Tripeptide Molecules
frequencies/cm-1
amide I
spectral
tripeptide molecules
solventsa
CCl4
CHCl3
CCl4
CHCl3
CCl4
water
CCl4
CHCl3
CCl4
water
ethyl acetate
water
typea
amide A
amide II
1
2
E
I
III
I
I
I
I
I
I
I
I
I
II
II
3364, 3320
3301
3301
3279
3281
3279
3280
3287
3286
3287
3281
3284
3277
3277
1674, 1663
1631
1647, 1631
1635
1536, 1516
1542
1559, 1533
1545
3
1634
1633
1634
1637
1637
1637
1634
1634
1541
1547
1544
1539
1538
1538
1543
1544
4
5
7
8
CHCl3
CCl4
1686, 1628
1689, 1628
1537
1535
a E denotes a specific mode except for a â-sheet structure (see text), and the others (I-III) are classified according to Figure 2. Measurement
temperature was 25.0 ( 0.5 °C.
Figure 6. FT-IR spectra of binary mixture and respective components (2 and 12) in CCl4 solution. Band split of 2 disappeared and a weak band
appeared at 1686 cm-1 (arrow at the bottom spectrum), which means the formation of an antiparallel â-sheet structure.
slighted despite their usefulness. Amyloid-like filaments whose
diameters varied from a few nanometers to 1 µm were depicted
in the AFM image. Figure 5 is a typical example of the AFM
image of the cast film from the CCl4 solution of 2, whose
secondary structure was a pseudoparallel â-sheet as previously
described. It is important that the similar outsides were obtained
using TEM and AFM, even if the samples were prepared by
the different procedures. The TEM sample was adsorbed fibers
onto an amorphous carbon film, whereas the AFM sample was
a dried specimen of an organogel on mica. We can observe
several branches in a tapping mode AFM image, whose widths
and height were 10-30 nm and 10-20 nm, respectively. The
most narrow branch should be an isolated fiber because the
diameter of the fiber evaluated from TEM images was 7-8 nm.
Although it was difficult to distinguish the respective â-sheet
structures from each other, namely, pseudoparallel- and parallel-
chain â-sheet structures, only from the AFM images, it is
noteworthy that a three-dimensional image of the â-sheet
assemblage can be visualized because such a three-dimensional
image has not yet been reported. The detailed morphological
aspect of the sub-µm AFM imaging is under experimental study.
The AFM and the TEM results showed that the tripeptide-
containing molecules formed a â-sheet assemblage. A similar
structure was observed within the cast films of amphiphiles
containing the other tripeptide residues. As previously de-
scribed, all of the amphiphiles except for 1 formed a parallel
â-sheet structure despite the large structural difference. How-
ever, the parallel â-sheet could be transformed into an antipar-
allel counterpart when the molecule was mixed with another
tripeptide-containing amphiphile 12 whose tripeptide part has
an opposite direction. For example, when the tripeptide-
containing amphiphile 2 was mixed with the single chain
amphiphile 12, a weak band at 1686 cm-1, which is evidence
of an antiparallel â-sheet, appeared together with a strong amide
I band at 1629 cm-1 similar to that of the tripeptide molecule
8 (Figure 6). The amphiphile 12 alone, of course, formed a
parallel â-sheet. On the other hand, it was possible to reform
the pseudoparallel â-sheet structure of 2 into a parallel â-sheet
structure. In this case, the pseudoparallel â-sheet structure of
2 transformed into a parallel â-sheet structure, on mixing with
a parallel â-sheet-forming 5 or 9. The transformations between
secondary structures are associated with the natural amyloid
formation, because it is believed that a segment of a protein in
the â-sheet form, especially a prion peptide, catalyzes the further
conversion of segments from an R-helix to the â-sheet form.1,15
(15) Gasset, M.; Baldwin, M. A.; Lloyd, D. H.; Gabriel, J.-M.; Holtzman,
D. M.; Cohen, F.; Fletterick, R.; Prusiner, S. B. Proc. Natl. Acad. Sci. U.S.A.
1992, 89, 10940-10944.

Amyloid-Like â-Sheet Assemblage
J. Am. Chem. Soc., Vol. 120, No. 47, 1998 12199
Conclusions
detection of the amide I bands. On the other hand, the
tripeptide-containing amphiphiles self-organized into a parallel
â-sheet assemblage, in which no H-bonding other than that
between the peptide linkages was present. Therefore, the present
assemblage should be a good model for the â-sheet polypeptides.
Morphological similarity also suggests that the fibrous model
including a parallel â-sheet structure should be helpful in
studying the structure and function of abnormal peptides such
as the prion and the amyloid.
There is controversy concerning the amide modes due to
parallel â-sheet conformation because of the model polypep-
tide’s lack of known X-ray structure for this conformation.12
Hence whether a â-sheet is parallel or antiparallel has not been
rigorously examined. Also, an interconversion between the two
structures has not been attempted. Here we demonstrated that
most of the tripeptide-containing amphiphiles formed a parallel
â-sheet structure in their assemblages. Emphasis is placed on
the controllability of the â-sheet structure within the assemblage.
The parallel â-sheet structure can be transformed into an
antiparallel counterpart by modifying the chemical structure,
changing the solvent, and mixing with other tripeptide molecules
possessing an opposite direction. It is sometimes difficult to
study amide modes when a tripeptide itself was used because
the intermolecular H-bonding between the amino group and the
carboxyl group appeared in the amide I region. The presence
of a protective group on the amino group also disturbs the
Acknowledgment. We thank Professor Yoshio Okahata,
Department of Biomolecular Engineering, Tokyo Institute of
Technology, for the AFM observations. This work was sup-
ported in part by a Grant-in-Aid for Scientific Research (No.
970018) from the Futaba Electronics Memorial Foundation,
Chiba, Japan.
JA981363Q