Cloning of the Endoglucanase Gene from a Bacillus amyloliquefaciens PSM 3.1 in Escherichia coli Revealed Catalytic Triad Residues Thr-His-Glu
- 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.
DOI: https://doi.org/10.3844/ajbbsp.2010.268.274
Copyright: © 2010 Zeily Nurachman, Sari Dewi Kurniasih, Ferra Puspitawati, Sarwono Hadi, Ocky Karna Radjasa and Dessy Natalia. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
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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