Screening interesting biocatalysts directly from soil samples is a more convenient and applicable approach than conventional cultivation-dependent ones. In our present work, a soil-derived metagenomic library containing 24,000 transformants was constructed with an efficient strategy for cloning xylanase genes. A gene encoding the enzyme (XynH) able to hydrolyze xylan was obtained. Similarity analysis revealed that this enzyme is a new member in the family 10 of xylanases. The molecular mass of XynH purified from Escherichia coli was estimated to be 39 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis. It was found to display the maximal activity at lower temperature, under weakly alkaline conditions, different from most of xylanases. The K m and Vmax values of XynH with birchwood xylan as substrate are 7.5 mg/ml and 190 μmol min−1 mg−1, respectively. It is greatly interesting to note that the activity of XynH was not reduced significantly by Mn2+, Zn2+, Co2+, Ag+, and Cu2+, even at the concentration of 5 mM, which strongly inhibits most of the other xylanases studied previously.
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We are very thankful to Prof. Li Yi in the NRS-Institute Armand-Frappier in Canada and Dr. James Sims in the University of Utah in USA for the refining of our manuscript. This research was supported by the National Natural Science Foundation of China (30600014 & 30770036), Wuhan City Science and Technology Plan (200760423162).
Yong Hu and Guimin Zhang contributed equally to this work.
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Hu, Y., Zhang, G., Li, A. et al. Cloning and enzymatic characterization of a xylanase gene from a soil-derived metagenomic library with an efficient approach. Appl Microbiol Biotechnol 80, 823 (2008). https://doi.org/10.1007/s00253-008-1636-6
- Soil microorganism DNA
- Metagenomic library construction
- Gene cloning
- Heterologous expression