Taxonomy of xylanolytic bacteria and characterization of xylanase from selected strains

Abstract

Seventy isolates of xylanase-producing bacteria were isolated from 45 samples of soil collected in Thailand. These bacteria were divided into sixteen groups based on their phenotypic and chemotaxonomic characteristics including 16s rRNA gene sequences of the representative strains. Sixty-one strains were Gram-positive rods belonged to Bacillus 25 isolates, Paenibacillus 24 isolates, Cohnella 4 isolates, Isoptericola and Jonesia each of 2 isolates, Microbacterium 3 isolates and Nocardioides 1 isolate. Each isolate of Gram-negative rods, was belonged to Acinetobacter, Aeromonas, Blastobacter, Ensifer, Pseudomonas, Sphingobacterium, Sphingomonas, Stenotrophomonas and Zobellella. They were identified as Bacillus licheniformis 4 isolates, Paenibacillus barengoltzii 3 isolates; each of 2 isolates was B. subtilis subsp. subtilis, B. niabensis, B. cereus, Isoptericola variabilis, Jonesia denitrificans and Microbacterium natoriense; and each of 1 isolate was B. nealsonii, P. macerans, P. timonensis, P. montaniterrae, P. dendritiformis, Nocardioides simplex, Acinetobacter junii, Aeromonas enteropelogenes, Ensifer adhaerens, Pseudomonas stutzeri, Stenotrophomonas maltophilia and Zobellella denitrificans. In addition, the novel species of Bacillus 2 isolates, Paenibacillus 12 isolates, Cohnella 4 isolates and each isolate of Microbacterium, Blastobacter, Sphingobacterium, Sphingomonas were identified based on the differential phenotypic, chemotaxonomic characteristics and 16S rRNA gene sequences similarity (96.0-98.8%). The Gram-positive rod-shaped, spore forming bacteria, Paenibacillus thailandensis sp. nov., P. nanensis sp. nov., P. xylanisolvens sp. nov., Cohnella thailandensis sp. nov, C. xylanilytica sp. nov. and C. terrae sp. nov. were proposed. Bacillus sp. P2-3 produced the highest xylanase activity, when compared to other isolates. Thus, P2-3 was selected for further study. Corn cob was found to be the most preferred substrate for xylanase production. After optimization of medium composition and pH, the yield was increased about 2 times when compared with the initial medium. The partially purified xylanase from P2-3 had molecular weight of 17.7 kDa. The enzyme had a maximal activity at 60°C and pH 6. Stability remained more than 50% at 30–40°C and pH 3-11. The xylanase activity was activiated by the addition of 1 mM Ca²⁺, Mg²⁺, Mn²⁺, DTT, and β-Me. In contrast, the xylanase activity was slightly inhibited by Fe²⁺, PMSF and SDS. The partially purified xylanase had the highest hydrolytic activity toward Oat spelt xylan, but no activity toward β-glucan, carboxymethylcellulose and pectin.