Purification and characteriation of a thermostable {beta}-xylosidase from thermoanaerobacter ethanolicus

Journal Article · Tue Sep 01 04:00:00 EDT 1992 · Journal of Bacteriology

OSTI ID:625672

Shao, W; Wiegel, J ^[1]

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1. Univ. of Georgia, Athens, GA (United States)

A highly thermostable {beta}-xylosidase, exhibiting similarly high activities for arylxylose and arylarabinose, was purified (72-fold) to gel electrophoretic from the ethanologenic thermophilic anaerobe Thermoanaerobacter ethanolicus. The isoelectric point is pH 4.6; the apparent molecular weight is around 165,000 for the native enzyme (gel filtration and gradient polyacrylamide gel electrophoresis) and 85,000 for the two subunits (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). The enzyme exhibited the highest affinity toward p-NO{sub 2}-phenyl xyloside (pNPX) (substrate concentration for half-maximal activity = 0.018 mM at 82{degrees}C and pH 5.0) but the highest specific activity with p-NO{sub 2}-phenylarabinofuranoside, t{sub ept, 5 min}, the temperature for the maximum initial activity in a 5-min assay of the purified enzyme, was observed around pH 5.9 and 93{degrees}C; however at 65 and 82{degrees}C (pH 5.0 to 5.5). The pH curves and temperature curves for arylxylosides as substrates differed significantly from those for arylarabinosides as substrates. An incubation for 3 h at 82{degrees}C in the absence of substrate reduced the activity to around 75%. At 86{degrees}C the half-life was around 15 min. With pNPX as the substrate, an Arrhenius energy of 69 kJ/mol was determined. The N-terminal sequence did not reveal a high similarity to those from other published enzyme sequences.

• DOE Contract Number: FG09-89ER14059
• OSTI ID: 625672
• Journal Information: Journal of Bacteriology, Journal Name: Journal of Bacteriology Journal Issue: 18 Vol. 174; ISSN JOBAAY; ISSN 0021-9193
• Country of Publication: United States
• Language: English

Related Subjects

55 BIOLOGY AND MEDICINE
BASIC STUDIES
AFFINITY
AMINO ACID SEQUENCE
ELECTROPHORESIS
ENZYME ACTIVITY
MOLECULAR WEIGHT
PH VALUE