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Purification and partial characterization of α-l-arabinofuranosidase produced byThermomonospora fusca

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Abstract

Thermomonospora fusca produced a relatively high level of α-l-arabinofuranosidase when growing on oat spelt xylan as the main carbon and energy source. The enzyme exhibited maximum relative activity (0.136 U/g protein) at pH 9.0 with 54 and 55% activity remaining at pH of 4.5 and 11.0, respectively. The apparentKm value for the crude α-l-arabinofuranosidase preparation was 180 µmol/L 4-nitrophenyl α-l-arabinofuranoside; thevlim value was the release of 40 µmol/L 4-nitrophenol per min. Enzyme activity was eluted as a single peak (HPLC gel filtration chromatography) corresponding to molar mass of ≈92 kDa. Native electrophoresis of crude cell lysate confirmed the presence of a single active intracellular α-l-arabinofuranosidase component. SDS-PAGE of this enzyme, developed as zymogram, did not demonstrate any activity; denaturing gel was stained and a protein band of relative molar mass of 46 kDa was revealed. Isoelectric focusing of a purified α-l-arabinofuranosidase yielded a single protein band for the corresponding activity zone with pI 7.9. The enzyme was purified approximately 21-fold the mean overall yield was about 16%.

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Abbreviations

IEF:

isoelectric focusing

NPA:

4-nitrophenyl α-l-arabinofuranoside

PAGE:

polyacrylamide gel electrophoresis

SDS:

sodium dodecyl sulfate

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Authors and Affiliations

  1. Biyoloji Bölümü, Fen-Edebiyat Fakültesi, Mersin Üniversitesi, 33342, Mersin, Turkey

    M. Tuncer

  2. John Tabor Laboratories, Department of Biological Sciences, University of Essex, CO4 3SQ, Colchester, UK

    A. S. Ball

Authors
  1. M. Tuncer
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  2. A. S. Ball
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Correspondence to M. Tuncer.

Additional information

The work was carried out in the frame of a postgraduate studentship provided for the first author byUniversity of Essex (UK); the first author also acknowledges the scholarship provided byMersin University (Turkey).

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Tuncer, M., Ball, A.S. Purification and partial characterization of α-l-arabinofuranosidase produced byThermomonospora fusca . Folia Microbiol 48, 168–172 (2003). https://doi.org/10.1007/BF02930950

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  • DOI: https://doi.org/10.1007/BF02930950

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