N-glycans from Paramecium bursaria chlorella virus MA-1D: Re-evaluation of the oligosaccharide common core structure

Abstract

Paramecium bursaria chlorella virus MA-1D is a chlorovirus that infects Chlorella variabilis strain NC64A, a symbiont of the protozoan Paramecium bursaria. MA-1D has a 339-kb genome encoding ca. 366 proteins and 11 tRNAs. Like other chloroviruses, its major capsid protein (MCP) is decorated with N-glycans, whose structures have been solved in this work by using nuclear magnetic (NMR) spectroscopy and MALDI-TOF mass spectrometry along with MS/MS experiments. This analysis identified three N-linked oligosaccharides that differ in the non-stoichiometric presence of three monosaccharides, with the largest oligosaccharide composed of eight residues organized in a highly branched fashion. The N-glycans described here share several features with those of the other chloroviruses except that they lack a distal xylose unit that was believed to be part of a conserved core region for all the chloroviruses. Examination of the MA-1D genome detected a gene with strong homology to the putative xylosyltransferase in the reference chlorovirus PBCV-1 and in virus NY-2A, albeit mutated with a premature stop codon. This discovery means that we need to reconsider the essential features of the common core glycan region in the chloroviruses.

The Astounding World of Glycans from Giant Viruses

“…The purification protocol used for PBCV-1 enabled the identification of the glycan structures of other chloroviruses (Figure ), classified either in the same group of viruses as PBCV-1 or in one of the other three chlorovirus groups (section ). ,− Of note, the chlorovirus NY-2A was predicted to have two MCPs with a high level of identity between them, and when this method was applied to NY-2A, both glycoproteins were extracted and the N -glycans derived from both proteins were studied together. Collectively, these structural studies established that the chlorovirus MCPs are always N -glycosylated and also revealed other interesting features.…”

“…Initially, this conserved oligosaccharide core consisted of five monosaccharide units: 179 the four highlighted in Figure 9 together with a so-called distal Xyl residue. However, this view has recently changed after determining the glycan structure of chlorovirus MA-1D, 193 a NC64A virus that produces three different N-glycans, all missing the distal Xyl unit. Accordingly, the conserved core region for all chloroviruses now consists of four sugar units (Figure 9).…”

“…Accordingly, the conserved core region for all chloroviruses now consists of four sugar units (Figure 9). 193 All chlorovirus N-glycans have a Rha unit linked at the third position of the hyperbranched Fuc, which is a semiconserved element because its absolute configuration is D for all the viruses within the NC64A and Osy groups, while it is L and always methylated at the O-3 position in the Pbi and SAG groups. 179 The substitution pattern of the conserved core region inclusive of the semiconserved Rha unit appears to be virus-specific, and it can be considered as a glycan-based molecular signature for each virus.…”

“…With regard to MA-1D, another chlorovirus in the NC64A group, like NY-2A and PBCV-1, in silico analysis suggested the presence of two MCPs (named 609L and 635L), 193 again with a very high level of homology between them and with the MCPs of both PBCV-1 and NY-2A. The two MA-1D MCPs were predicted to have the same, in terms of sequence and position, glycosylation sites as the other two viruses, and MALDI MS analysis confirmed three of them: 280 NIPG, 302 NTGT, and 406 NTAT.…”

ICBS 2021: Looking Toward the Next Decade of Chemical Biology

“…Combination of two alkyne-functionalized metabolic reporters, namely ManNAl and SiaNAl, allowed imaging of both sialin-dependent and sialin-independent incorporation pathways. , Cristina De Castro (University of Napoli) described their work on viral glycosylation of giant chloroviruses, on which they identified many original glycans with a new core structure, highly conserved between all chloroviruses. These glycans can be introduced on various peptidic sequences with no typical consensus. − Paramecium bursaria Chlorella virus 1 (PBCV-1) encodes several glycosyltransferases, some of which are conserved through other chloroviruses. ,, These glycosyltransferases might become useful tools for chemical glycobiology. Lipopolysaccharide of Alcaligenes faecalis shows promising properties as an adjuvant for vaccines.…”