Divergent synthesis of amino acid-linked O-GalNAc glycan core structures.

Abstract

O-GalNAc glycans, also known as mucin-type O-glycans, are primary constituents of mucins on various mucosal sites of the body and also ubiquitously expressed on cell surface and secreted proteins. They have crucial roles in a wide range of physiological and pathological processes, including tumor growth and progression. In addition, altered expression of O-GalNAc glycans is frequently observed during different disease states. Research dedicated to unraveling the structure-function relationships of O-GalNAc glycans has led to the discovery of disease biomarkers and diagnostic tools and the development of O-glycopeptide-based cancer vaccines. Many of these efforts require amino acid-linked O-GalNAc core structures as building blocks to assemble complex O-glycans and glycopeptides. There are eight core structures (cores one to eight), from which all mucin-type O-glycans are derived. In this protocol, we describe the first divergent synthesis of all eight cores from a versatile precursor in practical scales. The protocol involves (i) chemical synthesis of the orthogonally protected precursor (3 days) from commercially available materials, (ii) chemical synthesis of five unique glycosyl donors (1-2 days for each donor) and (iii) selective deprotection of the precursor and assembly of the eight cores (2-4 days for each core). The procedure can be adopted to prepare O-GalNAc cores linked to serine, threonine and tyrosine, which can then be utilized directly for solid-phase glycopeptide synthesis or chemoenzymatic synthesis of complex O-glycans. The procedure empowers researchers with fundamental organic chemistry skills to prepare gram scales of any desired O-GalNAc core(s) or all eight cores concurrently.