Conformational consequences of protein glycosylation: preparation of O-mannosyl serine and threonine building blocks, and their incorporation into glycopeptide sequences derived from alpha-dystroglycan.

With the goal to investigate the structural impact of O-mannosyl glycosylation on alpha-dystroglycan, a glycoprotein that has an important role in the extracellular organization of muscle, glycopeptides derived from its mucin-like sequence have been prepared by solid-phase peptide synthesis. Two approaches have been explored to obtain needed mannosylated serine and threonine building blocks. With the alpha-carboxyl group unprotected, and with tetraaceto-1-fluoro-alpha-D-mannose as the sugar donor, the desired alpha-O-mannosyl-Fmoc-Ser/Thr formed, along with mannosyl ester isomers and the species with mannose attached to both hydroxyl and carboxyl functions. Relevant mechanistic questions and stability issues were elucidated. Alternatively, building blocks were made with the alpha-carboxyl protected/activated as the pentafluorophenyl (Pfp) ester. Glycopeptides synthesized herein contained 5-9 residues, and featured one, two, and four consecutive Ser and/or Thr residues O-glycosylated with mannose. Circular dichroism (CD) spectra for Man-containing glycopeptides recorded in water show them to be not well ordered. For one of the alpha-dystroglycan-derived sequences, the comparative conformational consequences of glycosylation by either Man or GalNAc have been examined by CD and NMR, with both methods showing a more organized structure when GalNAc is present. (c) 2007 Wiley Periodicals, Inc.