Arabinose-5-phosphate isomerase

D-Ribono-1,4-lactone (1) readily condenses with acetone, under acidic catalysis with mineral acids or anhydrous copper sulfate, to give 2,3-0-isopropylidene-D-ribono-1,4-lactone (16a), which was employed for the synthesis of 5-deoxy and 5-0-substituted derivatives of D-ribono- 1,4-lactone and D-ribitol (24). Acid removal of the 1,3-dioxolane protecting group gave products having probable inhibitory activity of arabinose 5-phosphate isomerase (25). Other applications of 16a for the synthesis of natural products will be discussed later. 

HYDRATION WATER a-L-ARABINOFURANOSIDASE d-ARABINOKINASE ARABINONATE DEHYDRATASE ARABINOSE ISOMERASE ARABINOSE-5-PHOSPHATE ISOMERASE Arachidonate 5-lipoxygenase, 5-LIPOXYGENASE ARACHIDONYL-CoA SYNTHETASE Arene oxide intermediate,... 

D-Arabinose-5-phosphate isomerase, the first key enzyme in the synthesis of KDO, catalyzes the interconversion of D-ribulose-5-phosphate and D-arabinose-5-phosphate. This enzyme was briefly studied by Volk (18) and later by Lim and Cohen (19). Due to the instability of the enzyme, we have only purified this enzyme 100-fold. The reversible reaction is readily monitored by measuring the formation of the keto-sugar from the aldo-sugar by the method of Dische and Borenfreund (20). The K values for D-ribulose-5-phojsphate and D-arabinose-5-phosphate are 0.9 to 1.5 and 1 to 3 x 10 M, respectively. 

Figure 3. Plausible mechanism for the conversion of D-ribulose-5-phosphate isomerase to n-arabinose-5-phosphate by n-arabinose-5-phosphate isomerase. It is a least-motion model consistent with the mechanisms of similar enzymes (21-23).

TABLE I. Inhibition of D-Arabinose-5-phosphate Isomerase by Substrate Analogues... 

The data in Table III indicate that the enzyme KD0-8-phos-phate synthase is more difficult to inhibit with phosphorylated substrate analogues than is D-arabinose-5-phosphate isomerase. Non-phosphorylated substrate analogues (Tables II) were also tested as inhibitors or substrates of KDO-8-phosphate synthase but as with D-arabinose-5-phosphate isomerase, these analogues were... 

D-Glucose-6-phosphate dehydrogenase, D-gluconate-6-phosphate dehydrogenase, KDO-8-phosphate synthase, KDO-8-phosphate phosphatase and CMP-KDO synthetase have been purified to homogeneity and characterized. Munson, Rasmussen and Osborn (15) have partially purified one KDO transferase. D-Arabinose-5-phosphate isomerase is very unstable and has only been purified about 100 fold. D-Ribose-5-phosphate isomerase activity is approximately 20x that... 

The isolation and purification of the enzymes involved in KDO biosynthesis was necessary for the study of potential inhibitors of the individual reactions. A number of phosphorylated substrate analogues were synthesized and tested as inhibitors of D-arabinose-5-phosphate isomerase since this enzyme is the first direct reaction involved in KDO biosynthesis. (We have found no evidence that D-arabinose-5-phosphate is required in any other biosynthetic reaction in E. coli and Salmonella typhimurium it... 

The isolation of mutants in S. typhimurium by Rick and Osborn (11,12) and mutants in E. coli by Nishijima and Raetz (44) that accumulate the Lipid A precursor indicate that KDO synthesis and Lipid A synthesis are not coordinately controlled. The initial steps in the synthesis of the Lipid A precursor are totally unknown. The temperature sensitive mutants of E. coli isolated by Nishijima and Raetz (44) that are defective in phosphaditylglyc-erol phosphate synthesis at 42°C and accumulate the Lipid A precursors indicate that there is some relationship between the synthesis of phosphatidylglycerol and LPS. The reasons for the acu-cumulation of the Lipid A precursors in this E. coli mutant are not obvious. We have shown that CDP-diglyceride, one of the substrates for phosphatidylglycerol phosphate synthesis, is an inhibitor of D-arabinose-5-phosphate isomerase with an 1 value... 

These enzymes vary widely in secondary and tertiary structure.1273 Mannose-6-phosphate isomerase is a 45 kDa Zn2+-containing monomer. The larger 65 kDa L-fucose isomerase, which also acts on D-arabinose, is a hexameric Mn2+-dependent enzyme.1273 L-Arabinose isomerase of E. coli, which interconverts arabinose and L-ribulose, is a hexamer of 60-kDa subunits128 while the D-xylose isomerase of Streptomyces is a tetramer of 43-kDa subunits.129 The nonenzymatic counterpart of the isomerization catalyzed by the enzyme is the base-catalyzed Lobry deBruyn-Alberda van Ekenstein transformation (Eq. 13-25).130... 

Measurements of the crude specific activity (mmoles of product synthesized per minute per mg of protein in the supernatant after a 50,000 x g centrifugation) of the two isomerases in E. coli indicated that the conversion of D-ribulose-5-phosphate to D-ri-bose-5-phosphate was approximately 20- to 30-fold greater than the conversion of D-ribulose-5-phosphate to D-arabinose-5-phosphate. This rate of reaction strongly pulls the reaction substrate to D-ribose-5-phosphate, since the isomerase reaction at equilibrium strongly favors the formation of the aldo-sugar over the key intermediate D-ribulose-5-phosphate. 

L-arabinose isomerase, which interconverts L-arabinose and L-ribulose araB, L-ribulokinase, which uses ATP to phosphorylate L-ribulose at C-5 araD, L-ribulose 5-phosphate epimerase, which interconverts L-ribulose 5-phosphate and L-xylulose 5-phosphate talB, transaldolase and tktA, transketolase. 

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