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Molecular and biochemical characterization of a new alkaline active multidomain xylanase from alkaline wastewater sludge

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Abstract

A xylanase gene, xyn-b39, coding for a multidomain glycoside hydrolase (GH) family 10 protein was cloned from the genomic DNA of the alkaline wastewater sludge of a paper mill. Its deduced amino acid sequence of 1,481 residues included two carbohydrate-binding modules (CBM) of family CBM_4_9, one catalytic domain of GH 10, one family 9 CBM and three S-layer homology (SLH) domains. xyn-b39 was expressed heterologously in Escherichia coli, and the recombinant enzyme was purified and characterized. Xyn-b39 exhibited maximum activity at pH 7.0 and 60 °C, and remained highly active under alkaline conditions (more than 80 % activity at pH 9.0 and 40 % activity at pH 10.0). The enzyme was thermostable at 55 °C, retaining more than 90 % of the initial activity after 2 h pre-incubation. Xyn-b39 had wide substrate specificity and hydrolyzed soluble substrates (birchwood xylan, beechwood xylan, oat spelt xylan, wheat arabinoxylan) and insoluble substrates (oat spelt xylan and wheat arabinoxylan). Hydrolysis product analysis indicated that Xyn-b39 was an endo-type xylanase. The K m and V max values of Xyn-b39 for birchwood xylan were 1.01 mg/mL and 73.53 U/min/mg, respectively. At the charge of 10 U/g reed pulp for 1 h, Xyn-b39 significantly reduced the Kappa number (P < 0.05) with low consumption of chlorine dioxide alone.

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Acknowledgments

This research was supported by the National Science and Technology Support Program (2011BADB02) and the China Modern Agriculture Research System (CARS-42) and the National “948” project (2011-G7-4).

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Authors and Affiliations

  1. Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, People’s Republic of China

    Yanyu Zhao, Kun Meng, Huiying Luo, Huoqing Huang, Tiezheng Yuan & Bin Yao

  2. Department of Microbial Engineering, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, Beijing, 100081, People’s Republic of China

    Yanyu Zhao & Peilong Yang

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  1. Yanyu Zhao
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  2. Kun Meng
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  3. Huiying Luo
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  4. Huoqing Huang
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  5. Tiezheng Yuan
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  7. Bin Yao
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Correspondence to Peilong Yang or Bin Yao.

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Yanyu Zhao and Kun Meng contributed equally to this paper.

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11274_2012_1186_MOESM1_ESM.jpg

Fig. S1 Amino acid sequence alignment of Xyn-b39 (JN020645) with two GH 10 xylanases from Paenibacillus sp. W-61 (BAC45001.1) and Thermobacillus composti KWC4 (EGZ39961). (JPEG 182 kb)

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Zhao, Y., Meng, K., Luo, H. et al. Molecular and biochemical characterization of a new alkaline active multidomain xylanase from alkaline wastewater sludge. World J Microbiol Biotechnol 29, 327–334 (2013). https://doi.org/10.1007/s11274-012-1186-z

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  • DOI: https://doi.org/10.1007/s11274-012-1186-z

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