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Characterization of an extracellular thermophilic chitinase from Paenibacillus thermoaerophilus strain TC22-2b isolated from compost

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Abstract

Paenibacillus thermoaerophilus strain TC22-2b, a thermophilic bacterium with an optimum growth temperature of 50–55 °C isolated from compost (55 °C) in Japan, secreted a chitinase into culture medium in the presence of colloidal chitin. Adding glucose, lactose, mannose, or sucrose to culture medium decreased the amount of chitinase in culture supernatants. This chitinase was purified by ammonium sulfate precipitation, colloidal chitin adsorption, and ion exchange chromatography. The apparent molecular mass of this enzyme was approximately 48 kDa, and its N-terminal amino acid sequence was determined to be Ala–Val–Ser–Thr–Gly–Lys–Lys. The optimum temperature and pH for chitinase activity were 60 °C and pH 4, respectively. The chitinase retained 68 % of its initial activity after incubation at 50 °C for 2 h. Using p-nitrophenyl N,N′-diacetyl-β-d-chitobioside [pNP-(GlcNAc)2] as a substrate, the K m, V max, and k cat values for this enzyme were 1.4 mM, 0.058 mM min−1, and 9.6 s−1, respectively. Analysis of hydrolysis products showed that the chitinase digested N-acetyl-chitooligosaccharides in an endo manner. N-acetylglucosamine dimers were not degraded by the enzyme. When colloidal chitin was used as the substrate, N-acetylglucosamine dimers, -trimers, and -tetramers were detected as hydrolysis products. Thus, the thermophilic chitinase may prove useful for industrial applications in chitooligosaccharide production from chitin biomass.

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Acknowledgments

We thank Ms. Mai Kido and Mr. Mitsuhisa Watatani for their technical advice.

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Authors and Affiliations

  1. Department of Environmental Engineering for Symbiosis, Faculty of Engineering, Soka University, 1-236 Tangi-machi, Hachioji, Tokyo, 192-8577, Japan

    Junko Ueda & Norio Kurosawa

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  1. Junko Ueda
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  2. Norio Kurosawa
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Correspondence to Junko Ueda.

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Ueda, J., Kurosawa, N. Characterization of an extracellular thermophilic chitinase from Paenibacillus thermoaerophilus strain TC22-2b isolated from compost. World J Microbiol Biotechnol 31, 135–143 (2015). https://doi.org/10.1007/s11274-014-1754-5

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  • DOI: https://doi.org/10.1007/s11274-014-1754-5

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