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Volume 136, Issue 4

Isolation and characterization of an extremely thermostable D-xylose isomerase from HB 8 Free

Abstract

The extremely thermophilic organism possesses high activities of enzymes catalysing the degradation of xylans and metabolizing -xylose via the pentose phosphate pathway. The -xylose isomerase (-xylose ketol-isomerase, EC 5.3.1.5), an important enzyme of this process, is efficiently induced by its substrate -xylose, and, to a lesser extent, by related pentoses and some derivatives of -xylose. The -xylose isomerase from has been purified by anion-exchange chromatography, chromatography on -xylose agarose and gel filtration. A single band migrating according to an of 50000 was obtained by SDS-PAGE. An of 196000 for the native enzyme, determined by gel filtration and ultracentrifugation in a glycerol gradient, suggested that the -xylose isomerase is a homomeric tetramer. Arrhenius plots of the enzyme activity of the -xylose isomerase were linear up to a temperature of 85 °C. At 70 °C the enzyme was inactivated in the absence of divalent cations, with a half-life of 4 d, while in the presence of Mn or Co it remained fully active for at least 1 month. The enzyme had an isoelectric point at 4·4 and showed a broad optimum in the pH range from 5·5 to 8·5. No significant differences in the pH and temperature behaviour could be observed when -xylose was compared with -glucose as substrate. Different methods of immobilization of the enzyme to solid supports as well as inclusion into nylon beads were studied. Attachment of the enzyme to epoxy-activated agarose and its co-aggregation with bovine serum albumin gave immobilized preparations with the same stability as free enzyme supplemented with Mn or Co.

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© Society for General Microbiology, 1990
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1990-04-01
2025-03-18
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/content/journal/micro/10.1099/00221287-136-4-679
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Isolation and characterization of an extremely thermostable D-xylose isomerase from Thermus aquaticus HB 8
Microbiology 136, 679 (1990); https://doi.org/10.1099/00221287-136-4-679
/content/journal/micro/10.1099/00221287-136-4-679
/content/journal/micro/10.1099/00221287-136-4-679
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