Resolution-Chromatography-Information-Collision-Dissociation-Ions-Identify-Sequence-Oligosaccharides-Fact-Oligosaccharides-Disaccharides-Nonasaccharides-Sample-a

Fragmentation was shown to yield mostly reducing end sequence fragments (Z(i) and Y(i)), enabling primary sequence assignment. lacto-n-neotetraose or patterns were also detected giving specific linkage information. The reducing end core (GalGlcNAcbeta1-3GalNAcol or GalGlcNAcbeta1-3(GlcNAcbeta1-6)GalNAcol) could be deduced from the pronounced glycosidic C-3 cleavage and A(i) type cleavages of the reducing end GalNAcol, together with the non reducing end fragment from the loss of a single substituted GalNAcol. Substitution patterns on GlcNAc residues were also found, indicative for C-4 substitution (,2)A(i) - H(2)O cleavage) and disubstitution of C-3 and C-4 (Z(i)Z(i) cleavages). This kind of fragmentation can be used for assigning the mode of chain elongation (Galbeta1-34GlcNAcbeta1-) and identification of Lewis type antigens like Lewis ax and Lewis by on O-linked oligosaccharides. In essence, negative ion LC-MS(2) was able to generate extensive data for understanding the overall glycosylation pattern of a sample, especially when only a limited amount of material is available.

Characterisation of sucrose lactate and other oligosaccharides found in the Analysis of the raffinose family oligosaccharide pathway in pea seeds with Peterbauer T(1), Lahuta LB, Blöchl A, Mucha J, Jones DA, Hedley CL, Gòrecki RJ, Raffinose family oligosaccharides (RFOs) are synthesized by a set of galactosyltransferases, which sequentially add galactose units from galactinol to sucrose. The accumulation of RFOs was studied in maturing seeds of two pea accumulated stachyose as the predominant RFO, whereas verbascose, the next higher homolog of stachyose, was almost absent. In seeds of the line RRRbRb, a high level of verbascose was accumulated alongside with stachyose. The increase in verbascose in developing RRRbRb seeds was associated with galactinol-dependent verbascose synthase activity. In addition, a galactinol-independent enzyme activity was detected, which catalyzed transfer of a galactose residue from one stachyose molecule to another. The two enzyme activities synthesizing verbascose showed an optimum at pH 7. Human Milk Glycans were almost undetectable in SD1.

Maximum activity of stachyose synthase was about 4-fold higher in RRRbRb compared with SD1, whereas the activities of galactinol synthase and raffinose synthase were only about 1-fold higher in RRRbRb. The levels of galactinol synthase and stachyose synthase activity were reflected by steady-state levels of corresponding mRNAs. We suggest that the accumulation of verbascose in RRRbRb was controlled by a coordinated up-regulation of the last steps of verbascose biosynthesis.Elucidation of the structure of an alanine-lacking core tetrasaccharide trisphosphate from the lipopolysaccharide of Pseudomonas aeruginosa mutant H4.Lipopolysaccharide (LPS) of Pseudomonas aeruginosa rough mutant H4 was isolated by hot waterphenol extraction followed by a modified phenolchloroformpetroleum ether procedure. Upon SDSPAGE, the LPS showed a strong major band corresponding to the expected rough-type LPS. Additional faint high molecular-mass bands revealed that the O-chain was present, indicating that the H4 mutant is genetically unstable.

Mild acid hydrolysis of the LPS removed lipid A and released a phosphorylated core oligosaccharide that was purified by gel-permeation chromatography and high-performance anion-exchange liquid chromatography. The oligosaccharide contained two residues of L-glycero-D-manno-heptose (Hep) and one residue each of 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) and GalNAc. Upon matrix-assisted laser desorptionionization mass spectroscopy in the negative ion mode, the main fraction expressed a peak for the molecular ion [M-H]- at mz 161, which was compatible with a carbamoylated, trisphosphorylated tetrasaccharide. The structure was further investigated using one- and two-dimensional homonuclear and heteronuclear correlated NMR spectroscopy at pD 3 and, after borohydride reduction, at pD 9. The NMR data of the two phosphorylated tetrasaccharides recorded at different pD allowed determination of the positions of the three phosphate (P) groups and the carbamoyl group (Cm) thus establishing the following structure of the core oligosaccharide [equation see text] Two unusual structural features in the core oligosaccharide of P.