Identification and characterization of novel cellulolytic and hemicellulolytic genes and enzymes derived from German grassland soil metagenomes |
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Authors: | Heiko Nacke Martin Engelhaupt Silja Brady Christiane Fischer Janine Tautzt Rolf Daniel |
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Institution: | 1.Department of Genomic and Applied Microbiology, G?ttingen Genomics Laboratory, Institute of Microbiology and Genetics,Georg-August University G?ttingen,G?ttingen,Germany;2.Biology Faculty,University of Osnabrück,Osnabrück,Germany |
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Abstract: | Soil metagenomes represent an unlimited resource for the discovery of novel biocatalysts from soil microorganisms. Three large-inserts
metagenomic DNA libraries were constructed from different grassland soil samples and screened for genes conferring cellulase
or xylanase activity. Function-driven screening identified a novel cellulase-encoding gene (cel01) and two xylanase-encoding genes (xyn01 and xyn02). From sequence and protein domain analyses, Cel01 (831 amino acids) belongs to glycoside hydrolase family 9 whereas Xyn01
(170 amino acids) and Xyn02 (255 amino acids) are members of glycoside hydrolase family 11. Cel01 harbors a family 9 carbohydrate-binding
module, previously found only in xylanases. Both Xyn01 and Xyn02 were most active at 60°C with high activities from 4 to 10
and optimal at pH 7 (Xyn01) and pH 6 (Xyn02). The cellulase gene, cel01, was expressed in E. coli BL21 and the recombinant enzyme (91.9 kDa) was purified. Cel01 exhibited high activity with soluble cellulose substrates
containing β-1,4-linkages. Activity with microcrystalline cellulose was not detected. These data, together with the analysis
of the degradation profiles of carboxymethyl cellulose and barley glucan indicated that Cel01 is an endo 1,4-β-glucanase.
Cel01 showed optimal activity at 50°C and pH 7 being highly active from pH range 5 to 9 and possesses remarkable halotolerance. |
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