A novel endo-beta-1,3(4)-glucanase gene, cel16A, was cloned from the fungus Humicola insolens Y1. The 988-bp full-length gene encoded a 286-residue polypeptide consisting of a putative signal peptide of 20 residues and a catalytic domain belonging to glycosyl hydrolase family 16. It was successfully overexpressed in Pichia pastoris GS115. The purified recombinant Cel16A exhibited highest specific activity toward barley beta-glucan, followed by lichenan and laminarin, but not toward CMC-Na, birchwood xylan, Avicel and filter paper, indicating that Cel16A is an endo-beta-1,3(4)-glucanases. Recombinant Cel16A had a pH optimum at 5.5 and a temperature optimum at 55 degrees C with a specific activity of 693 U/mg toward barley beta-glucan. It exhibited good stability over pH 5.0-9.0 and at temperatures up to 50 degrees C, retaining over 80% maximum activity. The K-m and V-max values of Cel16A for barley beta-glucan were 0.91 mg ml(-1) and 1530 mol min(-1).mg(-1), respectively. All these favorable enzymatic properties of Cel16A make it a good candidate for applications in various industries. (C) 2017 Elsevier Inc. All rights reserved.