characterization of the gdp-d-mannose biosynthesis pathway in coxiella burnetii the initial steps for gdp-β-d-virenose biosynthesis描述gdp-d-mannose生物合成途径的伯纳特氏立克次氏体的初始步骤gdp-β-d-virenose生物合成.pdfVIP

Characterization of the GDP-D-Mannose Biosynthesis Pathway in Coxiella burnetii: The Initial Steps for GDP-b- D-Virenose Biosynthesis Craig T. Narasaki, Katja Mertens, James E. Samuel* Texas AM University Health Science Center, College of Medicine, College Station, Texas, United States of America Abstract Coxiella burnetii, the etiologic agent of human Q fever, is a Gram-negative and naturally obligate intracellular bacterium. The O-specific polysaccharide chain (O-PS) of the lipopolysaccharide (LPS) of C. burnetii is considered a heteropolymer of the two unusual sugars b-D-virenose and dihydrohydroxystreptose and mannose. We hypothesize that GDP-D-mannose is a metabolic intermediate to GDP-b-D-virenose. GDP-D-mannose is synthesized from fructose-6-phosphate in 3 successive reactions; Isomerization to mannose-6-phosphate catalyzed by a phosphomannose isomerase (PMI), followed by conversion to mannose-1-phosphate mediated by a phosphomannomutase (PMM) and addition of GDP by a GDP-mannose pyrophosphorylase (GMP). GDP-D-mannose is then likely converted to GDP-6-deoxy-D-lyxo-hex-4-ulopyranose (GDP-Sug), a virenose intermediate, by a GDP-mannose-4,6-dehydratase (GMD). To test the validity of this pathway in C. burnetii, three open reading frames (CBU0671, CBU0294 and CBU0689) annotated as bifunctional type II PMI, as PMM or GMD were functionally characterized by complementation of corresponding E. coli mutant strains and in enzymatic assays. CBU0671, failed to complement an Escherichia coli manA (PMM) mutant strain. However, complementation of an E. coli manC (GMP) mutant strain restored capsular polysaccharide biosynthesis. CBU0294 complemented a Pseudomonas aeruginosa algC (GMP) mutant strain and showed phosphoglucomutase activity (PGM) in a pgm E. coli mutant