Characterization of Escherichia coli D-arabinose 5-phosphate isomerase encoded by kpsF: implications for group 2 capsule biosynthesis.
In Escherichia coli, there are multiple paralogous copies of the enzyme API [A5P (D-arabinose 5-phosphate) isomerase], which catalyses the conversion of the pentose pathway intermediate Ru5P (D-ribulose 5-phosphate) into A5P. A5P is a precursor of Kdo (3-deoxy-D-manno-octulosonate), an integral carbohydrate component of various glycolipids coating the surface of the OM (outer membrane) of Gram-negative bacteria, including LPS (lipopolysaccharide) and many group 2 K-antigen capsules. The K-antigen-specific API KpsF has been cloned from the uropathogenic E. coli strain CFT073 and its biochemical properties characterized. Purified recombinant KpsF [K-API (K-antigen API)] is tetrameric and has optimal activity at pH 7.8. The enzyme is specific for A5P and Ru5P, with K(m) (app) values of 0.57 mM for A5P and 0.3 mM for Ru5P. The apparent kcat in the A5P to Ru5P direction is 15 and 19 s(-1) in the Ru5P to A5P direction. While most of the properties are quite similar to its LPS API counterpart KdsD, the catalytic constant is nearly 10-fold lower. K-API is now the second Kdo biosynthetic related gene that has been characterized from the kps group 2 capsule cluster.
Meredith
Timothy C
TC
Department of Medicinal Chemistry, University of Michigan, Ann Arbor, MI 48109-1065, USA.
Woodard
Ronald W
RW
D019747:Aldose-Ketose Isomerases / Q000302:isolation & purification / Q000378:metabolism*; D016667:Bacterial Capsules / Q000096:biosynthesis*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000302:isolation & purification; D013379:Substrate Specificity
D029968:Escherichia coli Proteins; C102536:KpsF protein, E coli; D008670:Metals; D011994:Recombinant Proteins; D019747:Aldose-Ketose Isomerases; C041486:arabinose-5-phosphate isomerase