Recombinant Thermostable Thermomonospora fusca TF Endo-xylanase A and Its Immobilization on Modified Mesoporous SiO2 Microspheres for Manufacturing Xylooligosaccharides
Recombinant Thermostable Thermomonospora fusca TF Endo-xylanase A and Its Immobilization on Modified Mesoporous SiO2 Microspheres for Manufacturing Xylooligosaccharides
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DOI：
10.1007/s10562-017-1979-7
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发表时间：
2017-01
影响因子：
2.8
通讯作者：
Ming-qi Liu;X. Weng;Qian Wang;Wen-kang Huo;Xin Xu
中科院分区：
文献类型：
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作者：
Ming-qi Liu;X. Weng;Qian Wang;Wen-kang Huo;Xin Xu
Abstracttfxa(GenBank: U01242), which encoded the mature peptide ofThermomonospora fuscaTF xylanase A (TfxA), was high-level expressed inEscherichia coliBL21 (DE3). The recombinant xylanase, reETfxA, was both secreted into culture medium and in the cytoplasm. reETfxA showed high xylanase activity and was purified to homogeneity by Ni-affinity resin (182.8 and 681.4 U mg−1for crude and purified enzyme, respectively). Sodium dodecyl sulphate–polyacrylamide gel electrophoresis and Western blot analysis revealed that the molecular mass of reETfxA was approximately 42.7 kDa. The reETfxA was immobilized on a novel mesoporous SiO2microsphere (MSM)-coated chitosan via covalent bonds, which formed from Schiff base reaction between supports and enzyme. The binding capacity of the prepared MSM-coated chitosan particles to reETfxA was approximately 272.6 mg g−1-particles. The optimum temperature values of the free and immobilized reETfxA were 65 and 70 °C, respectively. Moreover, the optimum pH values of the free and immobilized reETfxA were pH 6.0 and 5.0, respectively. The reETfxA showed relatively high thermostability. When treated at 70 °C and pH 6.0 for 15 min, the residual activities of free and immobilized reETfxA were 84.2 and 100.2%, respectively. The results obtained from the HPLC analysis showed that immobilized reETfxA released xylooligosaccharides from oat spelt, beechwood and birchwood xylans, with xylotetraose, xylotriose, and xylopentaose as the major products, respectively. Additionally, the immobilized reETfxA could directly hydrolyze the wheat bran insoluble xylan.Graphical Abstract
