A Neurospora crassa beta-glucosidase with potential for lignocellulose hydrolysis shows strong glucose tolerance and stimulation by glucose and xylose

作者：Meleiro Luana Parras; Santos Salgado Jose Carlos; Maldonado Raquel Fonseca; Alponti Juliana Sanchez; Ribeiro Latorre Zimbardi Ana Lucia; Jorge Joao Atilio; Ward Richard John; Melo Furriel Rosa Prazeres

来源：Journal of Molecular Catalysis B: Enzymatic , 2015, 122: 131-140.

DOI：10.1016/j.molcatb.2015.09.003

• Neurospora crassa
• Recombinant beta-glucosidase
• Lignocellulose saccharification
• Glucose- and xylose-stimulation
• Simultaneous saccharification and fermentation

Product inhibition of beta-glucosidases is one of the principal factors limiting the efficiency of enzymatic lignocellulosic biomass hydrolysis, particularly at high-solids concentrations. The availability of beta-glucosidases with high catalytic efficiency for cellobiose hydrolysis at relatively low temperatures is of major importance for the development of efficient and cost-effective simultaneous saccharification and fermentation (SSF) processes. The gene encoding the beta-glucosidase from Neurospora crassa (gh1-1) was cloned and expressed in soluble form in Escherichia coli. The recombinant enzyme (GH1-1) was monomeric (54.2 kDa) and showed optimal temperature and pH of 40-45 degrees C and 5.5-6.5, respectively. Moreover, activities around 70% of the maximal were maintained at pH 5.0 and 35 degrees C. The enzyme was highly stable at pH 5.5-8.0 and 35 degrees C, and showed a half-life of 70min at 40 degrees C in water. GH1-1 showed similar apparent affinities for cellobiose (0.21 +/- 0.01 mmol L-1) and p-nitrophenyl-beta-D-glucopyranoside (pNP-Glc) (0.28 +/- 0.01 mmol L-1) but hydrolyzed cellobiose with 3.2-fold higher maximal velocity (52.0 +/- 3.1 U mg(-1)) and 4.3-fold higher catalytic efficiency (223.8 L mmol(-1) s(-1)). Hydrolysis of pNP-Glc by GH1-1 was maximally stimulated 1.8 and 2.0-fold by glucose and xylose at 100 and 150 mmol L-1 concentration, respectively. Moreover, the enzyme was tolerant to glucose up to 950 mmol L-1 and xylose up to 910 mmol L-1. Xylose (60-80 mmol L-1) also stimulated the cellobiase activity of GH1-1 about 3.6-fold. Altogether, the characteristics of GH1-1 reveal its excellent potential for application as a component of enzymatic cocktails for the hydrolysis of lignocellulosic biomass, both in separate hydrolysis and fermentation and SSF processes.

• 出版日期2015-12