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Biochim Biophys Acta Gen Subj ; 1864(7): 129601, 2020 07.
Article in English | MEDLINE | ID: mdl-32179131

ABSTRACT

BACKGROUND: Escherichia coli cells contain a homolog of presumed 5-keto-4-deoxyuronate isomerase (KduI) from pectin-degrading soil bacteria, but the catalytic activity of the E. coli protein (o-KduI) was never demonstrated. METHODS: The known three-dimensional structure of E. coli o-KduI was compared with the available structures of sugar-converting enzymes. Based on the results of this analysis, sugar isomerization activity of recombinant o-KduI was tested against a panel of D-sugars and their derivatives. RESULTS: The three-dimensional structure of o-KduI exhibits a close similarity with Pyrococcus furiosus cupin-type phosphoglucose isomerase. In accordance with this similarity, o-KduI was found to catalyze interconversion of glucose-6-phosphate and fructose-6-phosphate and, less efficiently, conversion of glucuronate to fructuronate. o-KduI was hexameric in crystals but represented a mixture of inactive hexamers and active dimers in solution and contained a tightly bound Zn2+ ion. Dilution, substrate binding and Zn2+ removal shifted the hexamer ⇆ dimer equilibrium to the dimers. CONCLUSIONS: Our findings identify o-KduI as a novel phosphosugar isomerase in E. coli, whose activity may be regulated by changes in oligomeric structure. GENERAL SIGNIFICANCE: More than 5700 protein sequences are annotated as KduI, but their enzymatic activity has not been directly demonstrated. E. coli o-KduI is the first characterized member of this group, and its enzymatic activity was found to be different from the predicted activity.


Subject(s)
Aldose-Ketose Isomerases/genetics , Glucose-6-Phosphate Isomerase/genetics , Protein Conformation , Aldose-Ketose Isomerases/ultrastructure , Amino Acid Sequence/genetics , Carbohydrate Metabolism/genetics , Catalysis , Crystallography, X-Ray , Escherichia coli/enzymology , Fructosephosphates/genetics , Glucose-6-Phosphate/genetics , Glucose-6-Phosphate Isomerase/ultrastructure , Pyrococcus furiosus/enzymology
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