Research Article Open Access

Cloning of the Endoglucanase Gene from a Bacillus amyloliquefaciens PSM 3.1 in Escherichia coli Revealed Catalytic Triad Residues Thr-His-Glu

Zeily Nurachman1, Sari Dewi Kurniasih1, Ferra Puspitawati1, Sarwono Hadi1, Ocky Karna Radjasa2 and Dessy Natalia1
  • 1 Institute Technology Bandung, Indonesia
  • 2 Diponegoro University, Indonesia

Abstract

Problem statement:  An Indonesian marine bacterial isolate, Bacillus amyloliquefaciens PSM 3.1 was isolated for hydrolyzing cellulose. A 1500-bp nucleotide fragment was amplified from the chromosomal DNA by the use of primers directed against the conserved sequence of Bacilli endoglucanase genes obtained from GenBank. Approach: The fragment was cloned and expressed in Escherichia coli. Results: The endoglucanase gene (eglII gene) had an open reading frame of 1500 nucleotides encoding a protein of 499 amino acids. The EglII protein belonged to Glycosyl Hydrolase family 5 (GH5) with a Cellulose Binding Module 3 (CBM 3). The structure model of the EglII protein revealed that the catalytic residues seemed to be Glu169 (as proton donor) and Glu257 (as nucleophile) and the catalytic triad residues were Thr256, His229 and Glu169. The EglII endoglucanase exhibited an optimum pH of 6.0 and temperature of 50°C and the enzyme tolerated to high salt concentration. Conclusion/Recommendations: This EglII endoglucanase is a promising candidate for many applications in biomass degradation.

American Journal of Biochemistry and Biotechnology
Volume 6 No. 4, 2010, 268-274

DOI: https://doi.org/10.3844/ajbbsp.2010.268.274

Submitted On: 28 October 2010 Published On: 31 December 2010

How to Cite: Nurachman, Z., Kurniasih, S. D., Puspitawati, F., Hadi, S., Radjasa, O. K. & Natalia, D. (2010). Cloning of the Endoglucanase Gene from a Bacillus amyloliquefaciens PSM 3.1 in Escherichia coli Revealed Catalytic Triad Residues Thr-His-Glu. American Journal of Biochemistry and Biotechnology, 6(4), 268-274. https://doi.org/10.3844/ajbbsp.2010.268.274

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Keywords

  • Bacillus amyloliquefaciens
  • Escherichia coli
  • catalytic triad residues
  • endoglucanase
  • cellobiohydrolases
  • cellulolytic
  • Galaxea sp
  • nucleotides
  • hydrolysis
  • agarose gel electrophoresis
  • synthesized, polysaccharide
  • DNA polymerase
  • carboxymethyl cellulose