Overexpression, purification and characterization of the Aspergillus niger endoglucanase, EglA, in Pichia pastoris

D. H X Quay, Farah Diba Abu Bakar, A. Rabu, M. Said, R. M. Illias, N. M. Mahadi, Osman Hassan, Abdul Munir Abd. Murad

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

Cellulases are industrially important hydrolytic enzymes applicable in the bioconversion of cellulosic biomass to simple sugars. In this work, an endoglucanase from Aspergillus niger ATCC 10574, EglA, was expressed in the methylotrophic yeast Pichia pastoris and the properties of the recombinant protein were characterized. The full length cDNA of eglA has been cloned into a pPICZαC expression vector and expressed extracellularly as a ~30 kDa recombinant protein in P. pastoris X-33. Pure EglA displayed optimum activity at 50°C and was stable between 30 and 55°C. The pH stability of this enzyme was shown to be in the range of pH 2+.0 to 7.0 and optimum at pH 4.0. EglA showed the highest affinity toward β-glucan followed by carboxymethyl cellulose (CMC) with a specific activity of 63.83 and 9.47 U/mg, respectively. Very low or no detectable hydrolysis of cellobiose, laminarin, filter paper and avicel were observed. Metal ions such as Mn2+, Co2+, Zn2+, Mg2+, Ba2+, Fe2+, Ca2+ and K+ showed significant augmentation of endoglucanase activity, with manganese ions causing the highest increase in activity to about 2+.7 fold when compared with the control assay, whereas Pd2+, Cu2+, SDS and EDTA showed inhibition of EglA activity.

Original languageEnglish
Pages (from-to)2101-2111
Number of pages11
JournalAfrican Journal of Biotechnology
Volume10
Issue number11
Publication statusPublished - Mar 2011

Fingerprint

Pichia pastoris
Pichia
Aspergillus niger
Cellulase
endo-1,4-beta-glucanase
Recombinant Proteins
recombinant proteins
Ions
enzyme stability
Cellobiose
Enzyme Stability
Cellulases
Carboxymethylcellulose Sodium
cellobiose
cellulases
Glucans
carboxymethylcellulose
glucans
Manganese
biotransformation

Keywords

  • Aspergillus niger
  • Cellulase
  • Endoglucanase
  • Pichia pastoris
  • Recombinant

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology
  • Genetics
  • Molecular Biology
  • Agronomy and Crop Science

Cite this

Overexpression, purification and characterization of the Aspergillus niger endoglucanase, EglA, in Pichia pastoris. / Quay, D. H X; Abu Bakar, Farah Diba; Rabu, A.; Said, M.; Illias, R. M.; Mahadi, N. M.; Hassan, Osman; Abd. Murad, Abdul Munir.

In: African Journal of Biotechnology, Vol. 10, No. 11, 03.2011, p. 2101-2111.

Research output: Contribution to journalArticle

Quay, D. H X ; Abu Bakar, Farah Diba ; Rabu, A. ; Said, M. ; Illias, R. M. ; Mahadi, N. M. ; Hassan, Osman ; Abd. Murad, Abdul Munir. / Overexpression, purification and characterization of the Aspergillus niger endoglucanase, EglA, in Pichia pastoris. In: African Journal of Biotechnology. 2011 ; Vol. 10, No. 11. pp. 2101-2111.
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