PMID:12718525
| Citation |
Howe, DL, Sundaram, AK, Wu, J, Gatti, DL and Woodard, RW (2003) Mechanistic insight into 3-deoxy-D-manno-octulosonate-8-phosphate synthase and 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase utilizing phosphorylated monosaccharide analogues. Biochemistry 42:4843-54 |
|---|---|
| Abstract |
Escherichia coli 3-deoxy-D-manno-octulosonate 8-phosphate (KDO8-P) synthase is able to utilize the five-carbon phosphorylated monosaccharide, 2-deoxyribose 5-phosphate (2dR5P), as an alternate substrate, but not D-ribose 5-phosphate (R5P) nor the four carbon analogue D-erythrose 4-phosphate (E4P). However, E. coli KDO8-P synthase in the presence of either R5P or E4P catalyzes the rapid consumption of approximately 1 mol of PEP per active site, after which consumption of PEP slows to a negligible but measurable rate. The mechanism of this abortive utilization of PEP was investigated using [2,3-(13)C(2)]-PEP and [3-F]-PEP, and the reaction products were determined by (13)C, (31)P, and (19)F NMR to be pyruvate, phosphate, and 2-phosphoglyceric acid (2-PGA). The formation of pyruvate and 2-PGA suggests that the reaction catalyzed by KDO8-P synthase may be initiated via a nucleophilic attack to PEP by a water molecule. In experiments in which the homologous enzyme, 3-deoxy-D-arabino-heptulosonate 7-phosphate (DAH7-P) synthase was incubated with D,L-glyceraldehyde 3-phosphate (G3P) and [2,3-(13)C(2)]-PEP, pyruvate and phosphate were the predominant species formed, suggesting that the reaction catalyzed by DAH7-P synthase starts with a nucleophilic attack by water onto PEP as observed in E. coli KDO8-P synthase. |
| Links |
PubMed Online version:10.1021/bi026553z |
| Keywords |
3-Deoxy-7-Phosphoheptulonate Synthase; Aldehyde-Lyases/chemistry; Aldehyde-Lyases/metabolism; Binding Sites; Carbohydrate Conformation; Escherichia coli/enzymology; Kinetics; Magnetic Resonance Spectroscopy; Monosaccharides/chemistry; Monosaccharides/metabolism; Phosphorylation; Protein Conformation; Recombinant Proteins/chemistry; Recombinant Proteins/metabolism; Substrate Specificity |
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