Steven vimr free#

Our results indicate that, unlike UDP-GlcNAc 2-epimerase, which promotes the biosynthesis of sialic acid, GlcNAc-6-P 2-epimerase plays a catabolic role. coli K92 is a ManNAc-inducible protein and is detected from the early logarithmic phase of growth. Genetic analysis revealed high homology with bacterial isomerases.

The 2-epimerase activity was activated by Na(+) and inhibited by mannose-6-P but not mannose-1-P. Under these conditions, the K(m) calculated for GlcNAc-6-P was 1.5 mM. The maximum activity was achieved at pH 7.8 and 37 degrees C. By SDS/PAGE the pure enzyme showed a molecular mass of 38.4 +/- 0.2 kDa. We have purified for the first time native GlcNAc-6-P 2-epimerase from bacterial source to apparent homogeneity (1 200 fold) using Butyl-agarose, DEAE-FPLC and Mannose-6-P-agarose chromatography. In prokaryotic organisms, UDP-N-acetylglucosamine (GlcNAc) 2-epimerase and GlcNAc-6-P 2-epimerase are two enzymes capable of generating ManNAc from UDP-GlcNAc and GlcNAc-6-P, respectively. Thus, the mechanisms that control intracellular ManNAc levels are important regulators of sialic acid production. N-Acetylmannosamine (ManNAc) is the first committed intermediate in sialic acid metabolism. This study describes the first use of NMR for analysing intermediate flux within the sialate biosynthetic pathway. Our results imply that the neuC gene product encodes an UDP-N-acetylglucosamine 2-epimerase that generates ManNAc directly from the dinucleotide-sugar precursor despite detection of only this enzyme's UDP-GlcNAc hydrolase activity. The combined results indicate that neither ManNAc-6-P nor specific or non-specific phosphatase are necessary to generate the requisite ManNAc for sialate biosynthesis. Direct biochemical analysis showed that ManNAc-6-P was stable in a nanE mutant extract. The product specificities for purified NanK and NanE were determined by chromatographic analyses. coli K1 derivatives engineered with different combinations of mutations in nanA (sialate aldolase or lyase), nanK (ManNAc kinase), nanE (ManNAc-6-P 2-epimerase), neuS (polysialyltransferase) and neuB (sialate synthase). This conclusion was supported by 31P NMR spectroscopy of E. Here we show that ManNAc-6-phosphate (ManNAc-6-P) is not an obligate sialate precursor in Escherichia coli K1.

Steven vimr free#

The first committed step in this process, the production of free N-acetylmannosamine (ManNAc), has not been defined. The metabolism of sialic acid thus is central to a range of host-pathogen interactions.

A variety of pathogens or commensals use at least one of four distinct mechanisms for decorating their surfaces with sialic acid as a strategy to avoid, subvert or inhibit host innate immunity.