Glycosyl Esters of Nucleoside Pyrophosphates

Kochetkov, N. K.; Shibaev, V. N.

doi:10.1016/s0065-2318(08)60385-6

Cited by 60 publications

(18 citation statements)

References 446 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…GalNAcA is thought to be synthesized in vivo via epimerization and dehydrogenation of UDP-GlcNAc, the main precursor of surface-associated carbohydrate synthesis (12,18,19). The product of the epimerization reaction, UDP-GalNAc, is an important intermediate for the synthesis of polysaccharide structures that contain GalNAcA or a derivative, not only in P. aeruginosa but also in other organisms.…”

mentioning

confidence: 99%

Expression, Purification, and Biochemical Characterization of WbpP, a New UDP-GlcNAc C4 Epimerase from Pseudomonas aeruginosa Serotype O6

Creuzenet¹,

Bélanger²,

Wakarchuk³

et al. 2000

Journal of Biological Chemistry

109

View full text Add to dashboard Cite

B-band lipopolysaccharide is an important virulence factor of the opportunistic pathogen Pseudomonas aeruginosa. WbpP is an enzyme essential for B-band lipopolysaccharide production in serotype O6. Sequence analysis suggests that it is involved in the formation of N-acetylgalacturonic acid. To test this hypothesis, overexpression and biochemical characterization of WbpP were performed. By using spectrophotometric assays and capillary electrophoresis, we show that WbpP is a UDP-GlcNAc C4 epimerase. The K m for UDPGlcNAc and UDP-GalNAc are 197 and 224 M, respectively. At equilibrium, 70% of UDP-GalNAc is converted to UDP-GlcNAc, whereas the yield of the reverse reaction is only 30%. The enzyme can also catalyze the interconversion of non-acetylated substrates, although the efficiency of catalysis is significantly lower. Only 15 and 40% of UDP-Glc and UDP-Gal, respectively, are converted at equilibrium. WbpP contains tightly bound NAD(H) and does not require additional cofactors for activity. It exists as a dimer in its native state. This paper is the first report of expression and characterization of a C4 UDP-GlcNAc epimerase at the biochemical level. Moreover, the characterization of the enzymatic function of WbpP will help clarify ambiguous surface carbohydrate biosynthetic pathways in P. aeruginosa and other organisms where homologues of WbpP exist.

show abstract

mentioning

confidence: 99%

Expression, Purification, and Biochemical Characterization of WbpP, a New UDP-GlcNAc C4 Epimerase from Pseudomonas aeruginosa Serotype O6

Creuzenet¹,

Bélanger²,

Wakarchuk³

et al. 2000

Journal of Biological Chemistry

109

View full text Add to dashboard Cite

show abstract

“…[1][2][3][4][5] They function as donors for glycosyl residues where a donor of glycosyl residues is required (i.e. glycogen biosynthesis, cellulose biosynthesis).…”

Section: Introductionmentioning

confidence: 99%

“…The glycogen metabolom, for instance, is highly dependent on the presence of monosaccharide nucleotides. Uridine-[5′]-diphospho- [1]-α-D-glucose (UDP-α-GLC) is the activated form of glucose and functions as donor of glucose in the biosynthesis of glycogen. Monosaccharide nucleotides are of great importance for the synthesis of cell walls of micro-organisms and plants.…”

Section: Introductionmentioning

confidence: 99%

“…6 Classical analysis uses different hydrolytic treatments which include alkaline hydrolysis or enzymatic cleavage with specific exoglycosidases and pyrophosphatases. 1 The obtained degradation products are then compared with authentic monosaccharide and glycosyl phosphate standards (thin layer chromatography, high-performance liquid chromatography, high-performance anion-exchange chromatography with pulsed amperometric detection, nuclear magnetic resonance etc.). 1,7 These methods require large amounts of sample, which are no easily obtainable from biological sources.…”

Section: Introductionmentioning

confidence: 99%

“…1 The obtained degradation products are then compared with authentic monosaccharide and glycosyl phosphate standards (thin layer chromatography, high-performance liquid chromatography, high-performance anion-exchange chromatography with pulsed amperometric detection, nuclear magnetic resonance etc.). 1,7 These methods require large amounts of sample, which are no easily obtainable from biological sources. Fast atom bombardment mass spectrometry (FABMS), [8][9][10][11] which was found to give good results in the detection and characterization of monosaccharide nucleotides, improved the analysis of this class of compounds.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Negative Ion Ultraviolet Matrix-Assisted Laser Desorption Ionization Mass Spectrometry and Post Source Decay of Glycosyl Esters of Nucleoside Pyrophosphates

Heinrich

Schäffer

Messner

et al. 2008

Eur J Mass Spectrom (Chichester)

View full text Add to dashboard Cite

Six different glycosyl esters of nucleoside pyrophosphates (monosaccharide nucleotides) were analyzed by means of matrix-assisted laser desorption/ionisation reflectron time-of-flight mass spectrometry (MALDI/RToF MS) in the negative ion mode. Several matrices were evaluated and 3-hydroxypicolinic acid as well as α-cyano-4-hydroxycinnamic acid (CHCA) turned out to be the matrices of choice applying the thin layer technique to obtain maximum sensitivity for deprotonated molecular ion detection and maximal fragmentation particular with CHCA. The determination of the molecular mass with a mass accuracy below 0.1% was feasible with sample amounts in the lower femtomole range applying a MALDI desk-top mass spectrometer. A further important refinement of this technique was the use of post source decay (PSD) fragment ion analysis with a curved field reflector (which means no stepping of the reflector voltage). Detailed structural information of the six selected monosaccharide nucleotides could be obtained with PSD and differences in the fragmentation pattern were used to distinguish them. This method (based on molecular mass and PSD fragment ion analysis) has been applied to verify the presence of a glycosyl ester of nucleoside pyrophosphate in samples from Saccharomyces cerevisiae. KeywordsMALDI; post source decay; mass spectrometry; carbohydrate biosynthesis; glycosyl ester of nucleoside pyrophosphate; negative ions; ToF

show abstract

Synthesis of Sugar Nucleotides

Öhrlein

Ernst

Hart

et al. 2000

Carbohydrates in Chemistry and Biology

View full text Add to dashboard Cite

Recent progress in molecular glycobiology has revealed the important biological roles of numerous glycoconjugates [ 11. In particular cell-surface carbohydrates play key roles in cellular communication processes via selective adhesion phenomena [2, 31. They guide e.g. leukocyte extravasation [4], the homing of lymphocytes and adhesion of myeloid cells to activated platelets and the endothelium [5][6][7]. Besides these interactions carbohydrates are involved in fertilization events, cell growth, and parasitic infections of viruses and bacteria 181. A very recent finding is the involvement of certain cell surface a-galactosides in hyperacute rejection reactions; this is of pharmaceutical interest in organ transplantation 191.This makes sugars worthwhile targets for pharmaceutical applications [ 10, 1 11. Although chemical synthesis of oligosaccharides has reached a mature stage [ 12-141 the overall synthetic sequences are still lengthy and cumbersome [ 151. Biologically active carbohydrates are, moreover, composed of various different monosaccharide units and are often only active if presented in a polyvalent or multiantennary array to their respective receptors [ 16, 171. These multiantennary heteropolymers, in particular, have resisted efficient chemical synthesis. Recently biocatalysts have been found to be valuable tools for complementing the classical synthetic approach [ 181.Nature has invented several ways of assembling complex oligosaccharides [ 11. The Leloir pathway is one of the preferred routes of mammalian systems. The Leloir enzymes-the glycosyl transferases-transfer a monosaccharide unit from a nucleotide-activated donor substrate to a growing oligosaccharide chain with rigorous regio-and stereoselectivity [19] (see Scheme 1). Exclusively one of the many OH groups of the acceptor is glycosylated, in either an a or p mode! Neither the donor nor the acceptor sugar needs protection from the highly homofunctional reactants-often OH groups only are present on the sugars. Thus, selective protecting group manipulations, a preponderant task in the classical, chemical oligosaccharide assemblage can be reduced to an absolute minimum.

show abstract

Glycosyl Esters of Nucleoside Pyrophosphates

Cited by 60 publications

References 446 publications

Expression, Purification, and Biochemical Characterization of WbpP, a New UDP-GlcNAc C4 Epimerase from Pseudomonas aeruginosa Serotype O6

Expression, Purification, and Biochemical Characterization of WbpP, a New UDP-GlcNAc C4 Epimerase from Pseudomonas aeruginosa Serotype O6

Negative Ion Ultraviolet Matrix-Assisted Laser Desorption Ionization Mass Spectrometry and Post Source Decay of Glycosyl Esters of Nucleoside Pyrophosphates

Synthesis of Sugar Nucleotides

Contact Info

Product

Resources

About