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Expression and characterization of the recombinant Trichoderma virens endochitinase Cht2

Md. Illias, Rosli and S. A. A., Al-Rashed and F. D. A., Bakar and M., Said and O., Hassan and A., Rabu and A. M. A., Murad (2010) Expression and characterization of the recombinant Trichoderma virens endochitinase Cht2. African Journal Of Microbiology Research, 4 (16). pp. 1758-1769. ISSN 1996-0808

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Abstract

An endochitinase, Cht2, from Trichoderma virens UKM1 was expressed in the methylotrophic yeast Pichia pastoris, and its biochemical properties were characterized. Both the cht2 gene and its cDNA have been cloned and sequenced, the endochitinase gene cht2 encodes 321 amino acids from an open reading frame comprised of an 1169 bp nucleotide sequence separated by three introns. Cht2 is predicted to be an extracellular enzyme due to the presence of a signal peptide of 20 amino acids. Cht2 cDNA was cloned into the pPICZaC expression vector under the regulation of a methanol-inducing a promoter and transformed into P. pastoris X33. Expression in P. pastoris showed that the recombinant Cht2 was secreted into the culture medium with a protein size of approximately 35 kDa when induced with 0.5% methanol. Biochemical characterization of the partially purified enzyme showed a specific enzyme activity of 1.34 U/mg towards colloidal chitin at a pH of 6.0 and at a temperature of 35° C. The enzyme showed optimal activity at this pH and temperature and also showed higher affinity toward colloidal chitin in comparison to glycol chitin. It is stable in the pH range of 5.0 - 7.0 and in the temperature range of 30 - 55° C, where it retained more than 70% of its residual activity.

Item Type:Article
Uncontrolled Keywords:endochitinase, trichodermavirens, recombinant, characterization
Subjects:Q Science
Divisions:Biosciences and Bioengineering (Formerly known)
ID Code:27062
Deposited By: Liza Porijo
Deposited On:24 Jul 2012 02:16
Last Modified:24 Jul 2012 02:52

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