Purification and Characterization of a New Family 45 Endoglucanase, STCE1, from Staphylotrichum coccosporum and Its Overproduction in Humicola insolens

Abstract

ABSTRACT In the detergent industry, fungal endoglucanases have been used to release microfibrils (defibrillation) from the surface of dyed cellulosic fabrics to enhance color brightness. Although endoglucanases for laundry use must have various properties, such as a neutral or alkaline optimum pH, resistance to anionic surfactants and oxidizing agents (main components in detergents), and high defibrillation activity, all-purpose endoglucanases have not been obtained yet. As a result of screening of endoglucanases, a new family 45 endoglucanase (family 45 glycoside hydrolase), designated STCE1, was obtained and purified to apparent homogeneity from the culture supernatant of Staphylotrichum coccosporum NBRC 31817. The molecular mass of STCE1 was 49 kDa. The optimum pH for the carboxymethyl cellulase activity of STCE1 was 6.0, and the optimum temperature was 60°C. STCE1 was highly resistant to an anionic surfactant and an oxidizing agent. Furthermore, the defibrillation activities on dyed cotton and lyocell fabrics of STCE1 were higher than those of the other representative endoglucanases tested. These results indicate that STCE1 is an all-purpose enzyme for laundry use. A gene encoding STCE1, designated the stce1 gene, was cloned from S. coccosporum , and the complete sequence was determined. STCE1 consisted of three distinct domains: an N-terminal catalytic domain (family 45), a linker domain, and a C-terminal carbohydrate-binding module (family 1). The amino acid sequences of the catalytic domain of STCE1 were phylogenetically close to those of the family 45 endoglucanases EGL3, EGL4, and EGV from a Humicola sp. Hence, the stce1 gene was transferred into Humicola insolens and expressed. As a result, extremely high levels (0.90 mg protein per ml of culture supernatant, 27% of the total proteins) of the recombinant STCE1 were secreted as a mature form in the culture supernatant.