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Publication Abstract from PubMed

N-linked glycans play important roles in various cellular and immunological events. Endo-beta-N-acetylglucosaminidase (ENGase) can release or transglycosylate N-glycans and is a promising tool for the chemoenzymatic synthesis of glycoproteins with homogenously modified glycans. The ability of ENGases to act on core-fucosylated glycans is a key factor determining their therapeutic utility because mammalian N-glycans are frequently alpha-1,6-fucosylated. Although the biochemistries and structures of various ENGases have been studied extensively, the structural basis for the recognition of the core fucose and the asparagine-linked GlcNAc is unclear. Herein, we determined the crystal structures of a core fucose-specific ENGase from the caterpillar fungus Cordyceps militaris (Endo-CoM), which belongs to glycoside hydrolase family 18. Structures complexed with fucose-containing ligands were determined at 1.75-2.35 A resolutions. The fucose moiety linked to GlcNAc is extensively recognized by protein residues in a round-shaped pocket, while the asparagine moiety linked to the GlcNAc is exposed to the solvent. The N-glycan-binding cleft of Endo-CoM is Y-shaped and that several lysine and arginine residues are present at its terminal regions. These structural features were consistent with the activity of Endo-CoM on fucose-containing glycans on rituximab (IgG) and its preference for a sialobiantennary substrate. Comparisons with other ENGases provided structural insights into their core fucose tolerance and specificity. In particular, Endo-F3, a known core fucose-specific ENGase, has a similar fucose-binding pocket, but the surrounding residues are not shared with Endo-CoM. Our study provides a foothold for protein engineering to develop enzymatic tools for the preparation of more effective therapeutic antibodies.

Structural basis for the specific cleavage of core-fucosylated N-glycans by endo-beta-N-acetylglucosaminidase from the fungus Cordyceps militaris.,Seki H, Huang Y, Arakawa T, Yamada C, Kinoshita T, Iwamoto S, Higuchi Y, Takegawa K, Fushinobu S J Biol Chem. 2019 Sep 23. pii: RA119.010842. doi: 10.1074/jbc.RA119.010842. PMID:31548313^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.