黄曲霉产β-1,4-木聚糖酶发酵条件的优化及其酶学特性

Optimization of Fermentation Conditions for β-1,4-xylanase Production by Aspergillus flavus and Its Enzymatic Properties

研究发现一株高产β-1,4-木聚糖酶的黄曲霉,拟优化其产酶条件并分析该酶的酶学特性。采用单因素法优化其摇瓶发酵条件,聚丙烯酰胺凝胶电泳(SDS-PAGE)结合酶谱法判定酶的特性以及薄层色谱分析法(TLC)判定酶的水解特异性。结果表明,产酶最佳培养基为:麸皮3.5%、磷酸氢二铵3.0%、吐温-60 1.0%、NaCl 0.5%、MgSO_4·7H_2O 0.05%和KH_2PO_4 0.075%;黄曲霉在35℃下培养5 d达到最高酶活力115.08 U/mL,为未优化时的9.4倍。该菌能分泌两种β-1,4-木聚糖酶,分子量约为20.1和31.0 kDa,均不同于已有报道。粗酶的最适pH和最适温度分别为MOPS 7.5和55℃,在pH5.5～9.0及30～50℃范围内能保持酶活力的稳定。该酶不仅能水解榉木木聚糖产生低聚木糖,还能微弱降解大麦葡聚糖,有助于它在木质纤维素降解领域中的应用。

This study was aimed at optimization of the fermentation conditions forβ-1,4-xylanase production by Aspergillus flavus and characterization of its enzymatic properties.The fermentation conditions in shake flask were optimized by a single-factor experiment.And the enzymatic properties of the enzyme were analyzed by SDS-PAGE,zymogram and TLC.The results indicated that the optimal culture medium components forβ-1,4-xylanase production were 3.5%wheat bran,3.0%(NH4)2HPO4,1.0%Tween-60,0.5%NaCl,0.05%MgSO4·7 H2O and 0.075%KH2PO4.The highest enzyme activity of 115.08 U/mL was obtained when the strain was cultured at 35℃for 5 d in the optimized culture medium,which was 9.4 times compared with the initial activity.Twoβ-1,4-xylanases were secreted by A.flavus,and their molecular mass were different from all previous studies.The crude enzyme’s optimal temperature and pH were determined to be 55℃and 7.5,respectively.It was stable at 30~50℃and within pH5.5~9.0.The enzyme could not only efficiently hydrolyze beechwood xylan to produce xylooligosaccharides,but also weakly degrade the barley glucan,suggesting it was a good candidate for lignocelluloses’degradation.