TY - JOUR T1 - A library of structurally homogeneous human <em>N</em>-glycans synthesized from microbial oligosaccharide precursors JF - bioRxiv DO - 10.1101/118216 SP - 118216 AU - Brian S. Hamilton AU - Joshua D. Wilson AU - Marina A. Shumakovich AU - Adam C. Fisher AU - James C. Brooks AU - Alyssa Pontes AU - Radnaa Naran AU - Christian Heiss AU - Chao Gao AU - Robert Kardish AU - Jamie Heimburg-Molinaro AU - Parastoo Azadi AU - Richard D. Cummings AU - Judith H. Merritt AU - Matthew P. DeLisa Y1 - 2017/01/01 UR - http://biorxiv.org/content/early/2017/03/18/118216.abstract N2 - Synthesis of homogenous glycans in quantitative yields represents a major bottleneck to the production of molecular tools for glycoscience, such as glycan microarrays, affinity resins, and reference standards. Here, we describe a combined biological/enzymatic method termed bioenzymatic synthesis that is capable of efficiently converting microbially-derived precursor oligosaccharides into structurally uniform human-type N-glycans. Unlike starting material obtained by chemical synthesis or direct isolation from natural sources, which can be time consuming and costly to generate, bioenzymatic synthesis involves precursors derived from renewable sources including wild-type Saccharomyces cerevisiae glycoproteins and lipid-linked oligosaccharides from glycoengineered Escherichia coli. Following deglycosylation of these biosynthetic precursors, the resulting microbial oligosaccharides are subjected to a greatly simplified purification scheme followed by structural remodeling using commercially available and recombinantly produced glycosyltransferases including key N- acetylglucosaminyltransferases (e.g., GnTI, GnTII, and GnTIV) involved in early remodeling of glycans in the mammalian glycosylation pathway. Using this approach, preparative quantities of hybrid and complex-type N-glycans including asymmetric multi-antennary structures were generated all without the need of a specialized skillset. Collectively, our results reveal bioenzymatic synthesis to be a user-friendly methodology for rapidly supplying homogeneous oligosaccharide structures that can be used to understand the human glycome and probe the biological roles of glycans in health and disease. ER -