PMID: PMC7108604-1-4

 

    Legend: Gene, Sites

Title : Characterization of glycans

Abstract :
  1. Since the majority of endo H-sensitive glycans carried one sialic acid residue , they could be assumed to represent predominantly hybrid-type species
  2. This was corroborated by high-pH anion-exchange chromatography (HPAEC) of the desialylated glycans (Figure 4A)
  3. Only species H3 (14% of radioactivity within this fraction) coeluted with an oligomannosidic standard oligosaccharide (OM9), whereas the elution volumes of the two major components, H1 and H2 (62% and 24%), did not correspond to those of oligomannosidic standards, but to two hybrid-type standard glycans, MIII and MIV (Geyer et al., 1984), respectively (for detailed structures, see Figure 4 caption)
  4. To further prove their identity, half the amount of endo H-sensitive compounds obtained was digested with α-mannosidase from jack beans
  5. Rechromatography of the resulting products by HPAEC demonstrated that two products were formed eluting at the positions of authentic ManGlcNAcOH and GalGlcNAcMan2GlcNAcOH (Figure 4B)
  6. Digestion of the second half of the sample with β-galactosidase (Figure 4C) resulted in a shift of the elution volumes of the two hybrid-type species corresponding to the loss of one galactosyl residue each whereas the elution position of the oligomannosidic glycan H3 remained unchanged
  7. Therefore, it may be assumed that the two major oligosaccharide alditols, H1 and H2, represent hybrid-type species with one N-acetyllacto-samine antenna and two or three a-linked mannosyl residues
  8. From their sensitivity towards endo H, it may be further concluded that the terminal mannosyl residue present in H1 glycans is a1,3-linked (Kobata, 1979)
  9. The minor component H3, on the other hand, represented an oligomannosidic glycan with nine Man residues
  10. Neutral complex type glycans, obtained after individual desialylation of isocharged species, were chromatographically characterized by HPAEC (Figure 5) and gel filtration using a Bio-Gel P-4 column (data not shown)
  11. Although determination of chromatographic parameters does not allow a structural assignment a priori, comparison with the elution volumes of a set of appropriate oligosaccharide standards in at least two different chromatographic systems gives reliable results (Liedtke )
  12. The glucose units obtained from internal calibration with isomaltooligosaccharides were compared with those of authentic fucosylated di-, two isomers of tri-, tetrα-, and bisected diantennary standard oligosaccharide alditols
  13. The majority of glycans thus identified (Figure 5A–D; fractions F01, F11 , F13, F21, F31, F32) represented fucosylated species with two, three, or four N-acetyllactosamine antennae in addition to a few species not coeluting with any of these standard glycans (fractions F02, F12 )
  14. This assumption was supported by gel filtration data which suggested the presence of incomplete triantennary species lacking one (or two) galactosyl residue(s)
  15. To further substantiate these assignments, the four fractions of complex type glycans as well as the respective oligosaccharide standards were treated with β-galactosidase from D.pneumoniae
  16. Resulting products were again analyzed by HPAEC and, in part, also by gel filtration
  17. As shown in Figure 5E–H, compounds identified as fucosylated di-, tri-, and tetraantennary species coeluted with the respective agalacto oligosaccharide standards after β-galactosidase treatment
  18. Species F02 and F12 , formerly eluting at 4.62 and 4.44 glucose units (Figure 5A,B), now coeluted with triantennary agalacto oligosaccharide standards (Figure 5E,F) and thus may be assumed to comprise fucosylated triantennary species with one (or two) incomplete antenna(e)
  19. Differentiation between the possible isomeric forms, however, was not possible
  20. The elution position of species with 2.9 glucose units (Figure 5A) did not change after digestion with β-galactosidase , indicating the presence of incomplete species without any galactosyl residue
  21. Because of their elution properties, it may be further concluded that they were also lacking, to a certain extent, outer GlcNAc residues
  22. Since these glycans represented only minor constituents, they were not further analyzed
  23. The structural conclusions, drawn from the analytical data described, are summarized in Table I
Output (sent_index, trigger, protein, sugar, site):
Output(Part-Of) (sent_index, protein, site):
*Output_Site_Fusion* (sent_index, protein, sugar, site):

 

 

Protein NCBI ID SENTENCE INDEX