Structure of very unusual N-Glycans Associated with the Major Capsid Protein of Chlorovirus PBCV-1: Nature can synthesize a different class of complex N-Glycans

The envelope protein Vp54 from the prototype chlorovirus, Paramecium bursaria chlorella virus (PBCV-1) comprises ~40% of total virus protein and presents four N-glycosylated sites. In depth chemical, spectroscopical, and spectrometrical analysis has disclosed the structure of four different N-linked oligosaccharides and the type of substitution at each glycosylation site. Vp54 glycosylation is unusual in many respects: i) a ?-glucose, and not an aminosugar, is connected to the protein, ii) N-linked glycans are not located in a typical N-X-(T/S) consensus site, iii) four glycoforms are present. The structure of four glycoforms share a common core region and differences are related to the non-stoichiometric presence of two monosaccharides. The more abundant species comprises nine neutral monosaccharide residues, organized in a highly branched fashion; among the most relevant features, a dimethylated rhamnose as capping residue of the main chain, a hyperbranched fucose unit, and two rhamnose residues with opposite absolute configuration deserve to be mentioned. This protein glycosylation is new and different from what so far reported in the three domains of life. Considering that Chloroviruses and other members of the family Phycodnaviridae have a long evolutionary history possibly dating back to the time when eukaryotes arose from prokaryotes, we suggest that the chlorovirus glycosylation pathway is ancient, possibly existing prior to the development of the ER and Golgi, and involves new and still unexplored mechanisms.