Chemo-enzymatic synthesis of a bioactive peptide containing a glutamine-linked oligosaccharide and its characterization

• Published in: Biochimica et biophysica acta Vol. 1526; no. 3; pp. 242 - 248
• Main Authors: Haneda, Katsuji, Inazu, Toshiyuki, Mizuno, Mamoru, Iguchi, Reiko, Tanabe, Hiromi, Fujimori, Kenya, Yamamoto, Kenji, Kumagai, Hidehiko, Tsumori, Keiko, Munekata, Eisuke
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 15.06.2001
• Subjects: Amidohydrolases; Amino Acid Sequence; Animals; Carbohydrate Sequence; Chemo-enzymatic synthesis; Endo-β- N-acetylglucosaminidase; Glutamine - chemistry; Glutamine-linked oligosaccharide; Glycosylation; Guinea Pigs; Ileum - drug effects; Models, Chemical; Molecular Sequence Data; Muscle Contraction; Muscle, Smooth - drug effects; Oligosaccharides - chemistry; Peptide; Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; Substance P; Substance P - analogs & derivatives; Substance P - chemical synthesis; Substance P - pharmacology; Asn, N, l-asparagine; SGP, hen egg yolk glycopeptide having a disialo biantennary complex-type oligosaccharide; Substance P; SP, substance P; ACE, angiotensin-converting enzyme; NeuAc, N-acetyl- d-neuraminic acid; PNGase, peptide- N 4-( N-acetyl-β- d-glucosaminyl) asparagine amidase; PNGase F, PNGase from Flavobacterium meningosepticum; Fmoc, 9-fluorenylmethyloxycarbonyl; GlcNAc, N-acetyl- d-glucosamine; HPLC, high-performance liquid chromatography; Mpt-MA, dimethylphosphinothioic mixed anhydride; Boc, t-butoxycarbonyl; Gln, Q, l-glutamine; Endo-M, endo-β- N-acetylglucosaminidase from Mucor hiemalis; STF, a disialo biantennary complex-type oligosaccharide, (NeuAc-Gal-GlcNAc-Man) 2-Man-GlcNAc 2; Endo-β- N-acetylglucosaminidase; Man, d-mannose; Chemo-enzymatic synthesis; Peptide; Glutamine-linked oligosaccharide; Gal, d-galactose; TFA, trifluoroacetic acid; SGN, glycosyl asparagine having a disialo biantennary complex-type oligosaccharide
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/S0304-4165(01)00135-0

A bioactive peptide containing a glutamine-linked oligosaccharide was chemo-enzymatically synthesized by use of the solid-phase method of peptide synthesis and the transglycosylation activity of endo-β- N-acetylglucosaminidase. Substance P, a neuropeptide, is an undecapeptide containing two l-glutamine residues. A substance P derivative with an N-acetyl- d-glucosamine residue attached to the fifth or sixth l-glutamine residue from the N-terminal region was chemically synthesized. A sialo complex-type oligosaccharide derived from a glycopeptide of hen egg yolk was added to the N-acetyl- d-glucosamine moiety of the substance P derivative using the transglycosylation activity of endo-β- N-acetylglucosaminidase from Mucor hiemalis, and a substance P derivative with a sialo complex-type oligosaccharide attached to the l-glutamine residue was synthesized. This glycosylated substance P was biologically active, although the activity was rather low, and stable against peptidase digestion. The oligosaccharide moiety attached to the l-glutamine residue of the peptide was not liberated by peptide- N 4-( N-acetyl-β- d-glucosaminyl) asparagine amidase F.