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Effect of the chitin binding domain deletion from Bacillus thuringiensis subsp. kurstaki chitinase Chi255 on its stability in Escherichia coli

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Abstract

Bacillus thuringiensis subsp. kurstaki BUPM255 secretes a chitobiosidase Chi255 having an expected molecular weight of 70.665 kDa. When the corresponding gene, chi255, was expressed in E. coli, the active form, extracted from the periplasmic fraction of E. coli/pBADchi255, was of about 54 kDa, which suggested that Chi255 was excessively degraded by the action of E. coli proteases. Therefore, in vitro progressive C-terminal Chi255 deleted derivatives were constructed in order to study their stability and their activity in E. coli. Interestingly, when the chitin binding domain (CBD) was deleted from Chi255, an active form (Chi2555Δ5) of expected size of about 60 kDa was extracted from the E. coli periplasmic fraction, without the observation of any proteolytic degradation. Compared to Chi255, Chi255Δ5 exhibited a higher chitinase activity on colloidal chitin. Both of the enzymes exhibit activities at broad pH and temperature ranges with maximal enzyme activities at pH 5 and pH 6 and at temperatures 50°C and 40°C, respectively for Chi255 and Chi255Δ5. Thus, it was concluded that the C-terminal deletion of Chi255 CBD might be a nice tool for avoiding the excessive chitinase degradation, observed in the native chitinase, and for improving its activity.

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Abbreviations

B.:

Bacillus

E.:

Escherichia

CBD:

Chitin binding domain

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Acknowledgments

This work was supported by grants from the Tunisian « Ministère de l’Enseignement Supérieur, de la Recherche Scientifique et de la Technologie (MESRST) » and from the Islamic Educational, Scientific and Cultural Organization (ISESCO).

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Authors and Affiliations

  1. Laboratory of Biopesticides, Centre of Biotechnology of Sfax, P.O. Box ‘K’ 3038, Sfax, Tunisia

    F. Driss, A. Baanannou, S. Rouis, I. Masmoudi, N. Zouari & S. Jaoua

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  1. F. Driss
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  2. A. Baanannou
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  3. S. Rouis
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  4. I. Masmoudi
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  5. N. Zouari
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  6. S. Jaoua
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Correspondence to S. Jaoua.

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Driss, F., Baanannou, A., Rouis, S. et al. Effect of the chitin binding domain deletion from Bacillus thuringiensis subsp. kurstaki chitinase Chi255 on its stability in Escherichia coli . Mol Biotechnol 36, 232–237 (2007). https://doi.org/10.1007/s12033-007-0030-4

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  • DOI: https://doi.org/10.1007/s12033-007-0030-4

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