Short Articles
Bull. Chem. Soc. Jpn. Vol. 83, No. 7, 799–801 (2010)
799
labeled peptides, which gave non-natural type mannose- or
glucose-attached ¡-helix peptides. In the latter method, a
galactosylated Fmoc-threonine or a mannosylated Fmoc-
threonine was introduced to the ¡-helical model peptides
during the peptide assembly to produce O-glycoside peptides,
respectively. The HPLC profiles of synthetic products showed
that the latter method afforded a more favorable yield with
better profiles of the desired products than the former method
(Supporting Information). Thus, we have decided to employ the
building block strategy for the construction of glycopeptide
libraries. Their diversity arises from the use of four different
glycosides (glucoside, galactoside, mannoside, and lactoside),
their position and number (one or two) in addition to the
backbone amino acid sequence. Four building blocks of O-
glycosylated Fmoc-Thr (glucoside, mannoside, galactoside,
and lactoside) were successfully prepared in good yields
using BF3OEt2-mediated glycosylation of per-acetylated sugar
(Supporting Information). These building blocks were then
efficiently incorporated into peptides by using the Fmoc-solid
phase syntheses to give glycopeptides (Figure 1). All glyco-
Fingerprint-Detection of Sugar-
Binding Proteins Generated
by Labeled Structured
Glycopeptides Arrays
Takayasu Kawasaki, Takafumi Ohyama,
Akiyoshi Hirata, and Kiyoshi Nokihara*
HiPep Laboratories, 486-46 Nakatsukasacho,
Kamigyou-ku, Kyoto 602-8158
Received January 15, 2010
E-mail: noki@hipep.jp
Based on a novel biochip-concept involving labeled
structured peptides and the “protein-fingerprint” method,
the construction of O-glycopeptide-array on a novel chip
material and their use for biodetection are described.
α-helix (1-27)
β-sheet (28-54)
β-loop (55-80)
Since high-throughput protein-detection and characterization
systems are urgently required in the post-genomic era, we have
been developing a practical biochip system using synthetic
structured peptide libraries.1-3 Recently several peptide arrays
were reported for screening of epitope or discovery of ligands
of proteins,4,5 the novel concept of our system involves
protein-protein interactions that are mimicked by labeled
peptide-protein interactions. Visualization of interactions is
not in a “one to one” manner, but as bar-code like patterns with
fluorescent intensities generating “protein fingerprints.” The
structured peptide array is a sensing element for protein-
structure discrimination. The use of a peptide array as a
“protein-chip” affords significant advantages for industrial
production and applications for practical manufacturing,
storage, and delivery, compared to arrays with antibodies or
recombinant proteins.6 It is known that several toxic proteins,
such as Ricinus communis agglutinin (RCA), cholera toxin,
staphylococcal enterotoxin B, and Pseudomonas aeruginosa
lectin (PA-I) recognize cell-surface carbohydrates of the host
cells.7,8 Therefore, the sensitive detection system of these
carbohydrate-binding species in addition to protein-structure
recognition will be an effective tool for rapid detection and
may contribute to protection against various toxin-related
diseases. Hence, our peptide libraries consisting of several
hundreds of ¡-helix, ¢-sheet, and ¢-loop peptides have been
expanded to incorporate glycosides.
Glycopeptides are more difficult to synthesize and purify
than the parent peptides, although as mimetics glycopeptides
are better than glycosides alone, which has been reported
previously.9 Two strategies have been employed to generate
glycopeptides, the post-modification method and the building
block strategy.10 In the former method, mannosylamine or
glucuronic acid have been coupled to the ¾-amino group of Lys
of an ¡-helical TAMRA (5,6-carboxytetramethylrhodamine)-
Figure 1. List of glycopeptides (26, 27, 53, 54, 79, and 80
are non glycolsylated peptides as controls) synthesized in
the present study; 1-27: ¡-helix; 28-54: ¢-sheet; 55-80:
¢-loop. TG: glucosylated Thr; TM: mannosylated Thr; TGa
:
galactosylated Thr; TL: lactosylated Thr. C-Termini of
peptides are amide.