The first committed step in the biosynthesis of sialic acid byEscherichia coliK1 does not involve a phosphorylatedN‐acetylmannosamine intermediate
| Autor: | Susan M. Steenbergen, Eric R. Vimr, Michael A. Ringenberg |
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| Rok vydání: | 2003 |
| Předmět: |
Magnetic Resonance Spectroscopy
Mutant medicine.disease_cause Microbiology chemistry.chemical_compound Bacterial Proteins Biosynthesis N-Acetylmannosamine Escherichia coli medicine Histidine Cloning Molecular Phosphorylation Molecular Biology biology Escherichia coli Proteins Aldolase A Oxo-Acid-Lyases Hexosamines Lyase Molecular biology N-Acetylneuraminic Acid Sialyltransferases Sialic acid Phosphotransferases (Alcohol Group Acceptor) chemistry Biochemistry Mutation biology.protein Carbohydrate Epimerases Carrier Proteins N-Acetylneuraminic acid |
| Zdroj: | Molecular Microbiology. 50:961-975 |
| ISSN: | 1365-2958 0950-382X |
| DOI: | 10.1046/j.1365-2958.2003.03741.x |
| Popis: | 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. The metabolism of sialic acid thus is central to a range of host-pathogen interactions. The first committed step in this process, the production of free N-acetylmannosamine (ManNAc), has not been defined. Here we show that ManNAc-6-phosphate (ManNAc-6-P) is not an obligate sialate precursor in Escherichia coli K1. This conclusion was supported by 31P NMR spectroscopy of E. 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). The product specificities for purified NanK and NanE were determined by chromatographic analyses. Direct biochemical analysis showed that ManNAc-6-P was stable in a nanE mutant extract. 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. 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. This study describes the first use of NMR for analysing intermediate flux within the sialate biosynthetic pathway. |
| Databáze: | OpenAIRE |
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