Mass Spectrometry-based Avian Glycomics: Phylogenic Expression of Specific Terminal Glycotopes
Date Issued
2009
Date
2009
Author(s)
Su, Tseng-Hsiung
Abstract
Abstracthylogenetic expression of Galα1-4Gal glycotope among the avians has recently been systematically investigated by lectin and western blotting. In short, it was shown that only the egg white glycoproteins of avian species belonging to Neoaves but not those of Ratitae and Galloanserae would express this glycotope. However, the precise structures of its glycan carriers have thus far been established only for the pigeon egg white and IgG. Making use of a precious collection of egg white samples from over 1,000 species in hand, a systematic avian egg white glycomic analysis by mass spectrometry was initiated, aiming to map more extensively the phylogenic expression of Galα1-4Gal, along with any other novel epitopes, as well as to define the linkages of terminal sialylation in relation to influenza virus infection. To this end, egg white samples from over 20 avian species representing major taxonomical divisions were processed to release both N- and O-glycans and profiled by MALDI-MS after permethylation. Tentative assignment of the avian glycomic constituents was supported by MS/MS sequencing based on complementary low and high energy collision induced dissociation, as well as multistage activation MSn to define the linkage of sulfation, terminal galactosylation and sialylation. Additional glycomic mapping at the MS2 level by the total ion mapping functionality on a linear ion trap was developed and applied to define the entire sub-glycome on which the Gal-Gal glycotope is carried. Moreover, gel based glycomic analysis of individual protein bands enables a quick assessment of the distribution of individual glycans among distinct egg white glycoproteins. ollectively, applications of the developed experimental glycomic workflow successfully confirmed that Galα1-4Gal is phylogenetically distributed among the avian egg whites. A wide diversity of hybrid and complex type N-glycan structures were structurally defined, along with the identification of terminal Galβ1-4Gal, diLacNAc, LacdiNAc and sulfation. In short, the avian egg white N-glycomic heterogeneity was shown to be mostly due to incomplete α-/β-galactosylation, with or without bisecting GlcNAc, sialylation, and sulfation. In addition, Gal-Gal capping was found to occur not only on the egg white N-glycans but also on O-glycans, albeit much more restrictedly. Sulfation on the avian egg white N-glycans was widely detected and shown to be 6-linked to the terminal Gal residue of a LacNAc antenna, or the GlcNAc residue of the Gal-Gal-GlcNAc sequence. NeuAc-sialylation on gull egg white glycoproteins was shown to be predominantly in α2,3 linkage, whereas those on gull, turkey, guineafowl, and peafowl IgG samples prevailed in α2,6 form. Finally, gel-based analysis of a selected few egg white samples demonstrated that a significant difference in glycosylation profiles may exist among individual glycoprotein carriers for at least one avian species, but can be rather similar for others.
Subjects
mass spectrometry
avian
glycotope
glycomics
File(s)![Thumbnail Image]()
Loading...
Name
ntu-98-R96b46009-1.pdf
Size
23.32 KB
Format
Adobe PDF
Checksum
(MD5):a224f3c2b8d87795fc5a6fb573539297