D-Xylose (D-glucose) isomerase from Arthrobacter strain N.R.R.L. B3728. Purification and properties.
D-Xylose (D-glucose) isomerase was purified to homogeneity in yields of approx. 1 g/kg of wet cells from a strain of Arthrobacter that produces it as about 10% of total soluble protein. It is a tetramer of identical 43,114 Da subunits containing a preponderance of acidic residues and no cysteine. Partial protein sequences were determined as a step to gene cloning. It requires Mg2+, Co2+ or Mn2+ for activity, Mg2+ being best; Ca2+ is an inhibitor, competitive with Mg2+. It is a good D-glucose isomerase with kcat. 1200 min-1 at pH 8 at 60 degrees C, which is higher than that of any other enzyme of this class. L-Arabinose, D-ribose and D-lyxose are poor substrates, with kcat. 78, 31 and 3.7 min-1 respectively at pH 8 at 30 degrees C, compared with 533 min-1 for D-xylose. Xylitol is a true competitive inhibitor for D-xylose (Ki 0.3 mM), but D-sorbitol shows mixed inhibition (Ki 6.5 mM). For D-fructose the pH optimum at 60 degrees C is 8, and at pH 7 the Arrhenius activation energy is 75 kJ/mol over the range 30-70 degrees C.
Smith
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Centre for Biotechnology, Imperial College of Science, Technology and Medicine, London, U.K.
Rangarajan
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Hartley
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D019747:Aldose-Ketose Isomerases; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001173:Arthrobacter / Q000201:enzymology*; D001419:Bacteria / Q000201:enzymology; D002238:Carbohydrate Epimerases / Q000302:isolation & purification* / Q000378:metabolism; D002413:Cations, Divalent; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D013379:Substrate Specificity; D014357:Trypsin
D000596:Amino Acids; D002413:Cations, Divalent; D046911:Macromolecular Substances; D010446:Peptide Fragments; D014357:Trypsin; D002238:Carbohydrate Epimerases; D019747:Aldose-Ketose Isomerases; C023298:xylose isomerase