An endo-beta-1,4-mannanase encoding gene, man5, was cloned from Bispora antennata CBS 126.38, which was isolated from a beech stump. The cDNA of man5 consists of 1,299 base pairs and encodes a 432-amino-acid protein with a theoretical molecular mass of 46.6 kDa. Deduced MAN5 exhibited the highest amino acid sequence identity of 58% to a beta-mannanase of glycoside hydrolase family 5 from Aspergillus aculeatus. Recombinant MAN5 was expressed in Pichia pastoris and purified to electrophoretic homogeneity. The specific activity of the final preparation towards locust bean gum was 289 U mg(-1). MAN5 showed optimal activity at pH 6.0 and 70 degrees C and had good adaptation and stability over a broad range of pH values. The enzyme showed more than 60% of peak activity at pH 3.0-8.0 and retained more than 80% of activity after incubation at 37 degrees C for 1 h in both acid and alkaline conditions (pH 4.0-11.0). The K-m and V-max values were 1.33 mg ml(-1) and 444 mu mol min(-1) mg-1 and 1.17 mg ml(-1) and 196 mu mol min(-1) mg(-1) for locust bean gum and konjac flour, respectively. Of all tested metal ions and chemical reagents, Co2+, Ni2+, and beta-mercaptoethanol enhanced the enzyme activity at 1 mM, whereas other chemicals had no effect on or partially inhibited the enzyme activity. MAN5 was highly resistant to acidic and neutral proteases (trypsin, alpha-chymotrypsin, collagenase, subtilisin A, and proteinase K). By virtue of the favorable properties of MAN5, it is possible to apply this enzyme in the paper and food industries.