Characterization of the Primary Structure and the Microheterogeneity of the Carbohydrate Chains of Porcine Blood-Group H Substance by 500-MHz 1H-NMR Spectroscopy

Abstract

In blood‐group H substance from pig stomach cell linings, carbohydrate structures are known to occur which are O‐glycosidically linked via GalNAc to Ser or Thr of the polypeptide backbone. They have in common the Gal(β1→3)GalNAc core unit, which bears one or more N‐acetyllactosamine branches. The latter are terminated either by Fuc in (α1→2) linkage or by GlcNAc in (α1→44) linkage to Gal. Both terminal sequences are considered to be porcine blood‐group H antigenic determinants. Employing 500‐MHz 1H‐NMR spectroscopy, the spectral features of the oligosaccharide‐alditols corresponding to these chains were established. Insight could be gained into the microheterogeneity displayed by some of the alditol fractions as to the distribution of the terminating Fuc and GlcNAc residues over the various branches. Such information can hardly be obtained using other approaches.

Typing of core and backbone domains of mucin-type oligosaccharides from human ovarian-cyst glycoproteins by 500-MHz 1H-NMR spectroscopy

“…sequence linked to C-6 of Gal3 stems from the chemical shift effects of the H-3 and H-5 signals of GalNAc-ol and on the H-4 of Gal3. These are in line with those observed earlier [14]. Therefore, the structure of 2B is proposed to be as follows…”

“…The H-1 signal of the additional GlcNAc found at 4.607 ppm, suggests that the additional GalGlcNAc unit is linked b(1-6) to Gal3. This was previously observed in [14]. The Gal3 residue is thus serving as a branching point in the structure.…”

Primary structure of neutral oligosaccharides derived from respiratory‐mucus glycoproteins of a patient suffering from bronchiectasis, determined by combination of 500‐MHz 1H‐NMR spectroscopy and quantitative sugar analysis

“…The positions of the structural-reporter groups of Gal3 and GlcNAc6 are highly dependent on substitutions on these sugars. The 'H-NMR spectra of fractions 7 , l la, 15a and 16a are identical to those observed earlier for oligosaccharide-alditols (6,11,15.1A and 15.2B) obtained from cystic fibrosis sputum [l]. The corresponding primary structures are summarized in Scheme 3.…”

“…A reverse-phase column was used for further separation. Fractions 8,10,11,15,16,17,21,22 and 23 were subfractionated on a pBondapak TM C18 column (30 x 0.39 cm internal diameter, particle size 10 pm), eluted with water at a flow rate of 1 ml/min at room temperature, and gave fractions 8a-d; IOa, b; Ila, b; 15a, b; 16a-d; 17a, b; 21a, b; 22a-c; 23a-crespectively (Fig. 4).…”

Mucin synthesis

“…The NMR parameters are listed in Table 6. The addition of a fiGlcNAc residue in the product is indicated by an extra N-acetyl signal and a new doublet at 4.540 ppm (J1,z = 7.9 Hz), characteristic for the H-1 of a fl6-linked GlcNAc [3]. Major changes are seen in the H-3 resonances of GalNAc (from 4.027 ppm to 3.982 ppm), as well as the H-4 resonances of Gal (from 4.1 17 ppm to 4.086 ppm), indicating that the GlcNAc is linked to the 6 position of the 3-linked Gal rather than GalNAc.…”