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D L Fernandes

Publications and source records attributed to D L Fernandes.

9 recordsLinked to original sources
PubMed1998

Identification of highly fucosylated N-linked oligosaccharides from the human parotid gland.

The glycosylation of a number of constituents of human saliva is known to modify its biological roles, such as its lubricating properties and binding of microbial flora. Gillece-Castro et al. [Gillece-Castro, B. L., Prakobphol, A., Burlingame, A. L., Leffler, H. & Fisher, S. J. (1991) J. Biol. Chem. 266, 17358-17368] have proposed that the major glycan on the salivary proline-rich glycoproteins is a trifucosylated biantennary sugar with one difucosylated and one unfucosylated antenna. Furthermore, they proposed that the non-fucosylated antenna mediated adherence to a peridontal pathogen, Fusobacterium nucleatum. The detailed structures and roles of other highly fucosylated glycans that co-exist in the parotid gland are not fully known. In view of the influence of outer-arm fucosylation on carbohydrate recognition processes in general, this paper reports the use of a combination of HPLC (normal and reversed phase), matrix-assisted laser-desorption/ionisation (MALDI) mass spectrometry and exoglycosidase digestions to dissect the detailed structures of the most abundant of these polyfucosylated glycans. For measurement of reversed-phase HPLC retention times, new calibration units were used which paralleled the glucose units used for normal-phase HPLC. These differed in that the difference in retention times were compared with those derived from a ladder of 2-aminobenzamide-labelled arabinose oligomers instead of the corresponding oligomers from partially hydrolysed dextran. Over sixty neutral sugars were identified from the parotid gland and many of these were additionally found substituted with sialic acid (both alpha2-3-linked and alpha2-6-linked) and sulphate. These glycans were mainly bi- and tri-antennary sugars with up to five and seven fucose residues respectively, containing fucose alpha1-3-linked to the outer-arm GlcNAc residues and alpha1-2-linked to the galactose. All fucosylated structures contained a core (alpha1-6-linked) fucose. The detailed structure of the trifucosylated biantennary glycan was confirmed, together with the structures of another 12 fucosylated biantennary glycans. Smaller amounts of hybrid and tetraantennary structures were also found and bisected glycans were shown to be constituents of parotid glycoproteins for the first time. Acidic glycans were mainly substituted with sialic acid. Most were monosialylated as the presence of fucose on the antennae was found to suppress the addition of extra sialic acid moieties. The possible functional significance of highly fucosylated N-glycans is discussed in relation to their modification of the availability of other non-reducing terminal monosaccharides for recognition processes.

Carbohydrate Conformation↗
PubMed1984

The analysis of coupling networks in a complex oligosaccharide mixture derived from the Fc region of rabbit immunoglobulin G using 1H-1H correlated NMR spectroscopy combined with double quantum NMR spectroscopy.

In glycoproteins, even for those containing a single glycosylation site, diversity is manifest in the occurrence of a family of structurally-related yet distinct oligosaccharides. To date this 'microheterogeneity' is universal in mammalian glycoproteins. A method is described, using 1H-1H correlated and double quantum nuclear magnetic resonance NMR spectroscopy, for the assignment of proton resonances within a mixture of complex-type oligosaccharides derived from the Fc region of rabbit immunoglobulin G. The ability to assign resonances in heterogeneous populations will be of importance in the chemical shift analysis of the 1H-NMR spectra of glycopeptides since these cannot generally be separated on the basis of their carbohydrate sequence. The resulting assignments will be necessary before conformational studies on glycopeptides using nuclear Overhauser effects can be made.

Animals↗
PubMed1984

Multiple-step relayed correlation spectroscopy: sequential resonance assignments in oligosaccharides.

A general property of the high-resolution proton NMR spectra of oligosaccharides is the appearance of low-field well-resolved resonances corresponding to the anomeric (H1) and H2 protons. The remaining skeletal protons resonate in the region 3-4 ppm, giving rise to an envelope of poorly resolved resonances. Assignments can be made from the H1 and H2 protons to their J-coupled neighbors (H2 and H3) within this main envelope by using 1H-1H correlated spectroscopy. However, the tight coupling (J congruent to delta) between further protons results in poor spectral dispersion with consequent assignment ambiguities. We describe here three-step two-dimensional relayed correlation spectroscopy and show how it can be used to correlate the resolved anomeric (H1) and H2 protons with remote (H4, H5) protons directly through a linear network of couplings using sequential magnetization transfer around the oligosaccharide rings. Resonance assignments are then obtained by inspection of cross-peaks that appear in well-resolved regions of the two-dimensional spectrum. This offers a general solution to the assignment problem in oligosaccharides and, importantly, these assignments will subsequently allow for the three-dimensional solution conformation to be determined by using one-dimensional and two-dimensional nuclear Overhauser experiments.

Animals↗
PubMed1983

The use of two-dimensional correlated spectroscopy to obtain new assignments in the high-resolution 1H nuclear magnetic resonance spectrum of the biantennary complex oligosaccharide isolated from human serum transferrin by hydrazinolysis.

The asialo biantennary complex type oligosaccharide from human serum transferrin was isolated by hydrazinolysis, a method which results in the quantitative release of the intact oligosaccharide free of all amino acids. The 1H-NMR chemical shifts of the previously assigned anomeric and H-2 protons from the peripheral residues of the glycopeptide are identical to the corresponding values for the reduced oligosaccharide. The chemical shift of GlcNAc-1 H-1 proton in the reduced oligosaccharide was assigned by selective deuteration. Proton J connectivities were determined using two-dimensional 1H-1H correlated high resolution NMR spectroscopy. Twelve new assignments were made within the central envelope of the NMR spectrum and a further six were tentatively proposed. The ability to assign proton resonances in this way should allow further conformational studies of the oligosaccharide using nuclear Overhauser effects between the relevant assigned protons on different saccharide residues (Homans, S.W., Dwek, R.A., Fernandes, D.L. and Rademacher, T.W. (1982) FEBS Lett. 150, 503-506).

Borohydrides↗
PubMed1982

Solution conformation of the biantennary N-linked oligosaccharide of human serotransferrin using 1H NMR nuclear Overhauser effect measurements.

The conformation in solution of the biantennary complex type oligosaccharide unit derived from human serotransferrin has been investigated using 1H-1H Nuclear Overhauser Effect (NOE) measurements at 300 MHz. From quantitation of the NOE, the alpha(1-3) antenna is shown to exist in a preferred solution conformation with respect to the mannosyl-chitobiose core. The flexibility of the alpha(1-6) arm, together with the absence of NOE data between this arm and the core, indicates that, in contrast to the alpha(1-3) arm the alpha(1-6) arm has no preferred conformation with respect to the core.

Carbohydrate Conformation↗
PubMed1982

A comparison of the effects of infection with Eimeria maxima and dietary restriction on weight gain, plasma metabolites and liver glycogen in the immature fowl, Gallus domesticus.

The effects of Eimeria maxima or restricted pair-feeding on weight gain, plasma concentrations of protein, glucose, free fatty acids (FFA) and uric acid and liver glycogen were compared in immature fowl. Food intake/kg body weight and weight gain decreased during the acute phase of infection (days 5-7) while weight loss was prolonged for an extra day compared with pair-fed birds. During recovery, food intake/kg body weight of infected birds was greater than that of non-infected controls but there was no evidence for an increase in growth rate compared with controls when body weight was considered. Growth rate of pair-fed birds was greater than that of infected birds during recovery, indicating their better use of ingested food. Liver glycogen and plasma protein concentration were decreased during the acute phase of infection but the concentrations of plasma glucose, free fatty acid (FFA) and uric acid were not affected. In pair-fed birds liver glycogen was depleted, concentrations of plasma glucose and uric acid decreased and FFA increased, and these changes persisted for the remainder of the experiment. The findings are similar to those in birds whose food has been withheld and were probably due to the pattern of food intake imposed by the experimental protocol. It is concluded that the metabolic differences between infected and pair-fed birds are of doubtful significance.

Animals↗
PubMedDate not supplied

Association of rheumatoid arthritis and primary osteoarthritis with changes in the glycosylation pattern of total serum IgG.

Rheumatoid arthritis (RA) is a widely prevalent (1-3%) chronic systemic disease thought to have an autoimmune component; both humoral and cellular mechanisms have been implicated. Primary osteoarthritis (OA) is considered to be distinct from rheumatoid arthritis, and here damage is thought to be secondary to cartilage degeneration. In rheumatoid arthritis, immune complexes are present that consist exclusively of immunoglobulin, implying that this is both the 'antibody' (rheumatoid factor [RF]) and the 'antigen' (most commonly IgG). Autoantigenic reactivity has been localized to the constant-region (C gamma 2) domains of IgG. There is no evidence for a polypeptide determinant but carbohydrate changes have been reported. We have therefore conducted a study, simultaneously in Oxford and Tokyo, to compare in detail the N-glycosylation pattern of serum IgG (Fig. 1) isolated from normal individuals and from patients with either primary osteoarthritis or rheumatoid arthritis. The results, which required an evaluation of the primary sequences of approximately 1,400 oligosaccharides from 46 IgG samples, indicate that: (1) IgG isolated from normal individuals, patients with RA and patients with OA contains different distributions of asparagine-linked bi-antennary complex-type oligosaccharide structures, (2) in neither disease is the IgG associated with novel oligosaccharide structures, but the observed differences are due to changes in the relative extent of galactosylation compared with normal individuals. This change results in a 'shift' in the population of IgG molecules towards those carrying complex oligosaccharides, one or both of whose arms terminate in N-acetylglucosamine. These two arthritides may therefore be glycosylation diseases, reflecting changes in the intracellular processing, or post-secretory degradation of N-linked oligosaccharides.

Arthritis, Rheumatoid↗