High-level expression of a novel Penicillium endo-1,3(4)-β-d-glucanase with high specific activity in Pichia pastoris

Abstract A novel endo-1,3(4)-β-d-glucanase gene (bgl16C1) from Penicillium pinophilum C1 was cloned and sequenced. The 945-bp full-length gene encoded a 315-residue polypeptide consisting of a putative signal peptide of 18 residues and a catalytic domain belonging to glycosyl hydrolase family 16. Th...
Ausführliche Beschreibung

Abstract A novel endo-1,3(4)-β-d-glucanase gene (bgl16C1) from Penicillium pinophilum C1 was cloned and sequenced. The 945-bp full-length gene encoded a 315-residue polypeptide consisting of a putative signal peptide of 18 residues and a catalytic domain belonging to glycosyl hydrolase family 16. The deduced amino acid sequence showed the highest identity (82%) with the putative endo-1,3(4)-β-glucanase from Talaromyces stipitatus ATCC 10500 and 60% identity with the characterized β-1,3(4)-glucanase from Paecilomyces sp. FLH30. The gene was successfully overexpressed in Pichia pastoris. Recombinant Bgl16C1 constituted 95% of total secreted proteins (2.61 g $ l^{−1} $) with activity of 28,721 U $ ml^{−1} $ in a 15-l fermentor. The purified recombinant Bgl16C1 had higher specific activity toward barley β-glucan (12,622 U $ mg^{−1} $) than all known glucanases and also showed activity against lichenan and laminarin. The enzyme was optimally active at pH 5.0 and 55°C and exhibited good stability over a broad acid and alkaline pH range (>85% activity at pH 3.0–7.0 and even 30% at pH 11.0). All these favorable enzymatic properties make it attractive for potential applications in various industries. Ausführliche Beschreibung