Glycoprotein In Vitro N-Glycan Processing Using Enzymes Expressed in .

Protein N-glycosylation is a common post-translational modification that plays significant roles on the structure, property, and function of glycoproteins. Due to N-glycan heterogeneity of naturally occurring glycoproteins, the functions of specific N-glycans on a particular glycoprotein are not always clear. Glycoprotein in vitro N-glycan engineering using purified recombinant enzymes is an attractive strategy to produce glycoproteins with homogeneous N-glycoforms to elucidate the specific functions of N-glycans and develop better glycoprotein therapeutics.

A Neoglycoprotein-Immobilized Fluorescent Magnetic Bead Suspension Multiplex Array for Galectin-Binding Studies.

Carbohydrate-protein conjugates have diverse applications. They have been used clinically as vaccines against bacterial infection and have been developed for high-throughput assays to elucidate the ligand specificities of glycan-binding proteins (GBPs) and antibodies. Here, we report an effective process that combines highly efficient chemoenzymatic synthesis of carbohydrates, production of carbohydrate-bovine serum albumin (glycan-BSA) conjugates using a squarate linker, and convenient immobilization of the resulting neoglycoproteins on carboxylate-coated fluorescent magnetic beads for the development of a suspension multiplex array platform.

Size-Controlled Chemoenzymatic Synthesis of Homogeneous Oligosaccharides of W Capsular Polysaccharide.

(Nm) serogroup W (NmW) is one of the six meningococcal serogroups that cause majority of invasive meningococcal diseases (IMD). Its capsular polysaccharide (CPS) is a virulence factor and is a key component in NmW CPS-protein conjugate vaccines. The current clinically used NmW CPS-protein conjugate vaccines are effective but the costs are high and the products are heterogeneous at both the CPS and the conjugate levels.

A Chemoenzymatic Synthon Strategy for Synthesizing -Acetyl Analogues of -Acetylated W Capsular Polysaccharide Oligosaccharides.

-Acetylated sialic acid has been found in the serogroup W (NmW) capsular polysaccharide (CPS) and is a required structural component of clinically used NmW CPS-based polysaccharide and polysaccharide-conjugate vaccines. The role of sialic acid -acetylation in NmW CPS, however, is not clearly understood. This is partially due to the lack of a precise control of the percentage and the location of -acetylation which is labile and susceptible to migration.

Recent progress in chemical synthesis of bacterial surface glycans.

With the continuing advancement of carbohydrate chemical synthesis, bacterial glycomes have become increasingly attractive and accessible synthetic targets. Although bacteria also produce carbohydrate-containing secondary metabolites, our review here will cover recent chemical synthetic efforts on bacterial surface glycans. The obtained compounds are excellent candidates for the development of improved structurally defined glycoconjugate vaccines to combat bacterial infections.

Enterococcus faecalis α1-2-mannosidase (EfMan-I): an efficient catalyst for glycoprotein N-glycan modification.

While multiple α 1-2-mannosidases are necessary for glycoprotein N-glycan maturation in vertebrates, a single bacterial α1-2-mannosidase can be sufficient to cleave all α1-2-linked mannose residues in host glycoprotein N-glycans. We report here the characterization and crystal structure of a new α1-2-mannosidase (EfMan-I) from Enterococcus faecalis, a Gram-positive opportunistic human pathogen. EfMan-I catalyzes the cleavage of α1-2-mannose from not only oligomannoses but also high-mannose-type N-glycans on glycoproteins.

Regioselective One-Pot Multienzyme (OPME) Chemoenzymatic Strategies for Systematic Synthesis of Sialyl Core 2 Glycans.

O-GalNAc glycans or mucin-type glycans are common protein post-translational modifications in eukaryotes. Core 2 O-GalNAc glycans are branched structures that are broadly distributed in glycoproteins and mucins of all types of cells. To better understand their biological roles, it is important to obtain structurally defined Core 2 O-GalNAc glycans.