Hemocytes and Plasma of the Eastern Oyster (Crassostrea virginica) Display a Diverse Repertoire of Sulfated and Blood Group A-modified N-Glycans*

Abstract

Background:The eastern oyster, an important estuarine species, is parasitized by a protozoan in a galectin-dependent manner. Results: A variety of paucimannosidic, hybrid, and complex neutral and acidic N-linked oligosaccharides was found. Conclusion:The oyster possesses a complex repertoire of glycans with some features reminiscent of vertebrates. Significance: The N-glycome of the eastern oyster correlates well with the specificity of its own galectin CvGal1.

The Galectin CvGal1 from the Eastern Oyster (Crassostrea virginica) Binds to Blood Group A Oligosaccharides on the Hemocyte Surface*

“…8B), the residual binding suggest that the high affinity monoclonal antibodies can still bind to remaining GalNAc residues, which are not bound by CvGal1. Consistent results were obtained by Western blotting (61).…”

“…In this study, we used multiple approaches to reveal that CvGal1 recognizes blood group A oligosaccharides, also shown in a parallel glycomic study (61) to be present on the oyster hemocyte glycans. Finally, by proteomic analysis, we identified ␤-integrin and dominin among the hemocyte glycoproteins that are recognized by CvGal1.…”

N‐glycomic profiling of a glucosidase II mutant of Dictyostelium discoideum by ‘‘off‐line’’ liquid chromatography and mass spectrometry

“…Due to the wide variability in the N-glycans of various protists and invertebrates there is no single analytical workflow, but glycomic analyses of each species type very often have to be tailored to suit the properties of the glycans present. Thus, whereas N-glycans of nematodes, cestodes or trichomonads are either neutral or zwitterionic (31)(32)(33) , the protein-linked oligosaccharides of insects can be neutral, acidic or zwitterionic (34)(35)(36) and the N-glycans of oysters are neutral or sulphated (37) ; in the case of trematodes apparently only neutral glycans have been reported (38,39) .…”

“…Although these classes are the same as in the oyster (37) , a different approach is used by us when analysing glycans from these different sources; while no solid-phase extraction was necessary prior to HPLC of oyster N-glycans, we needed to pre-fractionate neutral and anionic N-glycomic pools from the slime mould prior to MS and HPLC analysis (23) . This acts not only as a polishing step during the N-glycan purification protocol, which is not necessary in every case, but separates different types of glycans and so the type of solid-phase extraction varies.…”

“…Subsequent fractionation by HPLC is also important to uncover the complexity of invertebrate glycomes; here, we have used both RP-HPLC, which is very suitable for examining isomeric glycans with the same mass, and HIAX, for size/charge separation. In other cases, 'pure' NP-HPLC may also be appropriate, as in the case of the oyster glycome (37) , or a '2D'-combination of NP-and RP-HPLC as applied to Caenorhabditis and Trichomonas (33,45) in order to achieve almost complete separation into single structures; particularly the latter greatly simplifies interpretation of exoglycosidase digests and avoids co-fragmentation of other glycans in the same sample when performing MS/MS.…”