Semisynthetic glycoproteins: preparation of glycosylated ribonuclease S′ analogues

✍ Scribed by Barbara Scolaro; Laura Biondi; Fernando Filira; Raniero Rocchi

• Book ID: 103958441
• Publisher: Elsevier Science
• Year: 1994
• Weight: 514 KB
• Volume: 22
• Category: Article
• ISSN: 0923-1137
• DOI: 10.1016/0923-1137(94)90117-1

✦ Synopsis

Glycopeptide analogues of ribonuclease S-peptide, des 16-20 [(Gal /3)Thr3]-S-peptide and des 16-20 [(Gal /3)Thr6]-S-peptide, were synthesized by the solid-phase procedure based on the Fmoc chemistry. Reassociation of the synthetic glycopeptides with S-protein yielded enzymatically active, glycosylated RNase S' variants. The conformational properties of the S-peptide analogues in aqueous buffer and in the presence of trifluoroethanol were investigated by circular dichroism spectroscopy. The activation curves of the S-protein with varying amounts of the synthetic analogues, using C > p as the substrate, were determined and the dissociation constants (Kd), the free energies of binding (-AG), and the kinetic parameters (K m and k 2) for the RNase S' adducts were calculated and compared with the corresponding values obtained for the S-peptide. The properties of the semisynthetic, glycosylated enzymes agree with the results of previous investigations on the conformational and catalytic features of S-peptide analogues and related RNase S'.