Escherichia coli YrbI Is 3-Deoxy-d-manno-octulosonate 8-Phosphate Phosphatase

• Published in: The Journal of biological chemistry Vol. 278; no. 20; pp. 18117 - 18123
• Main Authors: Wu, Jing, Woodard, Ronald W
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 16-05-2003
• Subjects: Amino Acid Sequence; Cations; Cloning, Molecular; Electrophoresis, Gel, Two-Dimensional; Electrophoresis, Polyacrylamide Gel; Escherichia coli - metabolism; Hydrogen-Ion Concentration; Kinetics; Magnesium - metabolism; Models, Chemical; Molecular Sequence Data; Molecular Weight; Phosphoric Monoester Hydrolases - chemistry; Phosphoric Monoester Hydrolases - metabolism; Protein Binding; Protein Structure, Tertiary; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Sequence Homology, Amino Acid; Substrate Specificity
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M301983200

3-Deoxy- d - manno -octulosonate 8-phosphate (KDO 8-P) phosphatase, which catalyzes the hydrolysis of KDO 8-P to KDO and inorganic phosphate,
is the last enzyme in the KDO biosynthetic pathway for which the gene has not been identified. Wild-type KDO 8-P phosphatase
was purified from Escherichia coli B, and the N-terminal amino acid sequence matched a hypothetical protein encoded by the E. coli open reading frame, yrbI . The yrbI gene, which encodes for a protein of 188 amino acids, was cloned, and the gene product was overexpressed in E. coli . The recombinant enzyme is a tetramer and requires a divalent metal cofactor for activity. Optimal enzymatic activity is
observed at pH 5.5. The enzyme is highly specific for KDO 8-P with an apparent K
m of 75 μ m and a k
cat of 175 s −1 in the presence of 1 m m Mg 2+ . Amino acid sequence analysis indicates that KDO 8-P phosphatase is a member of the haloacid dehalogenase hydrolase superfamily.