Preparation of peptides which mimic glycosphingolipids by using phage peptide library and their modulation on beta-galactosidase activity.
We describe the use of a phage-displayed random pentadecamer peptide library for searching glycosphingolipid mimicking peptides. Two phage clones (AD-1 and AD-2) were selected by biopanning using monoclonal antibody AD117m, directed to lactotetraosylceramide (Lc4Cer). The amino acid sequences of the selected clones showed high homology (VPPXFXXXY) in 9-mer. Three phage clones were selected by using monoclonal antibody H11, directed to neolactotetraosylceramide (nLc4Cer), the linkage isomer of Lc4Cer, and the displayed amino acid sequences were compared. One of these peptides showed the same amino acid sequence as that of AD-2 except for one amino acid substitution. Pentadecamer, 9-mer and point mutated 9-mer peptides were synthesized on the basis of the displayed amino acid sequences. Binding activity of the peptides to the monoclonal antibodies or Ricinus communis lectin showed that 9-mer peptides are enough to mimic the epitope carbohydrate structure. Furthermore, six of the synthesized peptides inhibited Jack bean beta-galactosidase activity towards nLc4Cer at a high concentration of the enzyme, whereas at lower enzyme concentrations some peptides showed potent activation of the enzyme activity. This is the first report of carbohydrate mimicking peptides which modulate glycosidase activity.
Taki
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T
Department of Biochemistry, School of Medicine, Tokyo Medical and Dental University, Japan.
Ishikawa
D
D
Hamasaki
H
H
Handa
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S
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001435:Bacteriophages; D001483:Base Sequence; D002240:Carbohydrate Sequence; D002518:Ceramides / Q000737:chemistry* / Q000494:pharmacology; D016384:Consensus Sequence; D007887:Fabaceae / Q000201:enzymology; D006028:Glycosphingolipids / Q000737:chemistry* / Q000494:pharmacology; D007700:Kinetics; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D009842:Oligopeptides / Q000737:chemistry* / Q000494:pharmacology*; D019151:Peptide Library; D037121:Plant Lectins; D010946:Plants, Medicinal; D012276:Ricin / Q000737:chemistry / Q000494:pharmacology; D016415:Sequence Alignment; D001616:beta-Galactosidase / Q000037:antagonists & inhibitors*
D000911:Antibodies, Monoclonal; D002518:Ceramides; D006028:Glycosphingolipids; D009838:Oligodeoxyribonucleotides; D009842:Oligopeptides; D019151:Peptide Library; D037121:Plant Lectins; D012276:Ricin; D001616:beta-Galactosidase