The simultaneous synthesis of 40 different s-linked O-glycopeptides from human intestinal mucin and porcine submaxillary gland mucin in a manual multiple column peptide synthesizer using the new glycosylamino acid building blocks Na-Fmoc-Ser(Ac3-~-D-GalpNAc)-OPfp (3) and Na-Fmoc-Thr(Ac3-~-D-GalpNAc)-OPfp ( 4) is described.
The important role in biological systems played by the surface carbohydrates on glycoproteins and cell membranes in recognition and transport processes has increased the interest and demand for a variety of glycopeptides as model compounds 1. For the study of the biosynthesis2 of mucin type glycoproteins synthetic O-glycopeptides, which carry an a-linked GalNAc-sugar on serine or threonine, are required. Recently, Brockhausen3 has shown that the enzymatic synthesis of O-glycan core structures is controlled by the peptide moiety. The activity of the 133-Galactosyltransferase, which glycosylates the 3-OH position in the GalNAc unit, is dependent of the peptide sequence, the chain length, the attachment site of the GalNAc and the number of GalNAc-residues in the substrate. N-terminally acetylated glycopeptide amides were much better substrates than the corresponding molecules with free amino and carboxy termini especially in the case of partially purified enzymes. For a more detailed enzymatic investigation a large number of synthetic glycopeptides 4 is needed. The concept to synthesize O-glycopeptides by using glycosylated hydroxy amino acids as building blocks for solid phase synthesis has been used successfully several times5.
In this paper the application of the two new building blocks 3 and 4 is reported. These key compounds were obtained by esterification of the acids 1 and 26 using pentafluorophenol (Pfp-OH) and dicyclohexylcarbodiimide (DCCI) in dry ethyl acetate at 0ยฐC. After chromatographic purification on silica gel under dry conditions5d and precipitation from diethyl ether the activated ester 3 and 4 were both obtained in 75% yield as stable solid materials (-24% of the valuable starting material could be recovered from each separation). The active esters 3 and 4 were characterized by 1D-and 2D-1H-NMR spectroscopy and FAB-MS7.