Selection of purification condition for recombinant endoglucanase originated from goat rumen bacteria in Escherichia coli

Nguyen Thi Quy, Nguyen Hong Duong, Dao Trong Khoa, Nguyen Khanh Hoang Viet, Nguyen Khanh Hai, Truong Nam Hai, Do Thi Huyen
Author affiliations

Authors

  • Nguyen Thi Quy
  • Nguyen Hong Duong
  • Dao Trong Khoa
  • Nguyen Khanh Hoang Viet
  • Nguyen Khanh Hai
  • Truong Nam Hai
  • Do Thi Huyen

DOI:

https://doi.org/10.15625/0866-7160/v42n1.14455

Keywords:

Escherichia coli Rosetta2, Carboxymethylcellulose (CMC), clear zone (halo), protein purification, recombinant endoglucanase.

Abstract

Cellulase is an important enzyme that plays a role in cleaving β-1,4 glucoside on cellulose to release glucose, which is of economic value and can be applied in many different fields. The 1545 bp endoglucanase gene mined from goat rumen's bacterial metagenomic data was expressed in Escherichia coli Rosetta2. In this study, the recombinant endoglucanase was purified by his-tag affinity chromatography with differrent processes, such as using phosphate buffer with or without sodium cloride, pretreatment of samples with ammonium sulphate before supplying into affinity column, using various concentration of imidazole for washing... Finally the endoglucanse was sucessfully purified by his-tag affinity column using sodium chloride-free phosphate buffer of which 150 mM and 400 mM imidazole were used for washing and enzyme elution, respectively. The resulting enzyme showed its high purity of 99%. CMC plate assay confirmed that although less active than commercial cellulase (Sigma), the recombinant cellulase hydrolyzed CMC to form a clear zone (halo) around the well. The purified enzyme is capable of using as material for further analysis.

 

Downloads

Download data is not yet available.

Metrics

Metrics Loading ...

References

Amore A., Pepe O., Ventorino V., Birolo L., Giangrande C., Faraco V., 2012. Cloning and recombinant expression of a cellulase from the cellulolytic strain Streptomyces sp. G12 isolated from compost. Microb. Cell Factories, 11: 164.

Cano-Ramírez C., Santiago-Hernández A., Rivera-Orduña F.N., García-Huante Y., Zúñiga G., Hidalgo-Lara M.E., 2016. Expression, purification and characterization of an endoglucanase from Serratia proteamaculans CDBB-1961, isolated from the gut of Dendroctonus adjunctus (Coleoptera: Scolytinae). AMB Express6.

Chuan Wei K.S., Teoh T.C., Koshy P., Salmah I., Zainudin A., 2015. Cloning, expression and characterization of the endoglucanase gene from Bacillus subtilis UMC7 isolated from the gut of the indigenous termite Macrotermes malaccensis in Escherichia coli. Electron.J. Biotechnol., 18: 103–109.

Lynd L.R., Weimer P.J., van Zyl W.H., Pretorius I. S.,2002. Microbial cellulose utilization: fundamentals and biotechnology. Microbiol. Mol. Biol. Rev. MMBR, 66: 506–577.

Maki M., Leung K.T., Qin W., 2009. The prospects of cellulase-producing bacteria for the bioconversion of lignocellulosic biomass. Int. J. Biol. Sci., 5: 500–516.

Nguyen K.H.V., Nguyen T.T., Truong N.H., Do T.H., 2019. Application of bioinformatic tools for prediction of active pH and temperature stability of endoglucanases based on coding sequences from metagenomic DNA data. Biological Forum, 11: 14–20.

Nguyen T.Q., Duong T.H., Dang T.N.H., Le N.G., Le Q.G., Do T.H., Nguyen V.D., Le T.T.H., Truong N.H., 2018. Enhanced soluble expression and efective purification of recombinant human interleukin-11 by SUMO fusion in Escherichia coli. Indian Journal of Biotechnology, 17: 579–585.

Pandey S., Kushwah J., Tiwari R., Kumar R., Somvanshi V.S., Nain L., Saxena A.K., 2014. Cloning and expression of β-1, 4-endoglucanase gene from Bacillus subtilis isolated from soil long term irrigated with effluents of paper and pulp mill. Microbiol. Res., 169: 693–698.

Seneesrisakul K., Guralp S.A., Gulari E., Chavadej S., 2017. Escherichia coli expressing endoglucanase gene from Thai higher termite bacteria for enzymatic and microbial hydrolysis of cellulosic materials. Electron. J. Biotechnol., 27: 70–79.

Downloads

Published

19-03-2020

How to Cite

Quy, N. T., Duong, N. H., Khoa, D. T., Hoang Viet, N. K., Hai, N. K., Hai, T. N., & Huyen, D. T. (2020). Selection of purification condition for recombinant endoglucanase originated from goat rumen bacteria in <i> Escherichia coli </i>. Academia Journal of Biology, 42(1). https://doi.org/10.15625/0866-7160/v42n1.14455

Issue

Section

Articles

Most read articles by the same author(s)

1 2 3 > >>