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Functional expression and characterization of a chitinase from the marine archaeon Halobacterium salinarum CECT 395 in Escherichia coli

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Abstract

The HschiA1 gene of the archaeon Halobacterium salinarum CECT 395 was cloned and overexpressed as an active protein of 66.5 kDa in Escherichia coli. The protein called HsChiA1p has a modular structure consisting of a glycosyl hydrolase family 18 catalytic region, as well as a N-terminal family 5 carbohydrate-binding module and a polycystic kidney domain. The purified recombinant chitinase displayed optimum catalytic activity at pH 7.3 and 40 °C and showed high stability over broad pH (6–8.5) and temperature (25–45 °C) ranges. Protein activity was stimulated by the metal ions Mg+2, K+, and Ca+2 and strongly inhibited by Mn+2. HsChiA1p is salt-dependent with its highest activity in the presence of 1.5 M of NaCl, but retains 20 % of its activity in the absence of salt. The recombinant enzyme hydrolysed p-NP-(GlcNAc)3, p-NP-(GlcNAc), crystalline chitin, and colloidal chitin. From its sequence features and biochemical properties, it can be identified as an exo-acting enzyme with potential interest regarding the biodegradation of chitin waste or its bioconversion into biologically active products.

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Acknowledgments

This work was developed within the Sustainable Use Network of Environmental and Agrifood Resources REDUSO and was supported by Grant 10PXIB310278PR (Xunta de Galicia). G.-F., B. has a predoctoral fellowship from the University of Vigo, Spain.

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  1. Faculty of Biology, Department of Functional Biology and Health Sciences, University of Vigo, 36310, Vigo, Spain

    Belén García-Fraga, Abigaíl F. da Silva, Jacobo López-Seijas & Carmen Sieiro

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  1. Belén García-Fraga
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  2. Abigaíl F. da Silva
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  3. Jacobo López-Seijas
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  4. Carmen Sieiro
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Correspondence to Carmen Sieiro.

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García-Fraga, B., da Silva, A.F., López-Seijas, J. et al. Functional expression and characterization of a chitinase from the marine archaeon Halobacterium salinarum CECT 395 in Escherichia coli . Appl Microbiol Biotechnol 98, 2133–2143 (2014). https://doi.org/10.1007/s00253-013-5124-2

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  • DOI: https://doi.org/10.1007/s00253-013-5124-2

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