Purification and Characterization of a Protease-Resistant Cellulase from Aspergillus niger

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Abstract

An endo-β-1,4-glucanase (EC 3.2.1.4) was purified from a culture filtrate of Aspergillus niger IFO31125 by column chromatography through TSK-gel DEAE-3SW and TSK-gel DEAE-5PW, and by gel filtration through TSK-gel G2000SW by high performance liquid chromatography. The enzyme was estimated to have a molecular weight of about 40 kDa by both gel filtration and SDS-polyacrylamide gel electrophoresis, and appeared to consist of a monomeric protein. It contained 8.9% carbohydrate. The optimal pH for activity was 6.0-7.0,and the stable pH range was 5.0-10.0. The optimum temperature at pH 6.0 was around 70℃. The enzyme was very thermally stable and no loss of original activity was found on incubation at 60℃ for 2 h. The enzyme efficiently hydrolyzed carboxymethylcellulose and lichenan, but crystalline forms of cellulose, curdlan, laminarin, cellobiose, p-nitrophenyl-β-D-glucopyranoside and p-nitrophenyl-β-D-cellobioside were barely hydrolyzed. The activity of the enzyme was inhibited by Hg^<2+> and Cu^<2+> but was not affected by other inhibitors of thiol enzymes such as p-chloromercuribenzoate and N-ethylmaleimide. N-Bromosuccinimide showed a strong inhibitory effect, suggesting that a tryptophan residue is essential for the activity of the enzyme. The N-terminal amino acid sequence of the enzyme showed considerable homology to those of endo-β-1,4-glucanases from some other microorganisms, including Sclerotinia sclerotiorum and Schizophyllum commune. The enzyme had very strong protease-resistance, and showed no loss of activity when incubated with proteases such as Savinase at 40℃, even for 2 weeks.

Journal

  • J. Ferment. Bioeng.

    J. Ferment. Bioeng. 79(2), 125-130, 1995-02-25

    The Society for Biotechnology, Japan

References:  28

Cited by:  6

Codes

  • NII Article ID (NAID)
    110002681976
  • NII NACSIS-CAT ID (NCID)
    AA10702688
  • Text Lang
    ENG
  • Article Type
    Journal Article
  • ISSN
    0922338X
  • Data Source
    CJP  CJPref  NII-ELS  Crossref 
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