Purification and Characterization of a Novel Fungal α-Glucosidase fromMortierella alliaceawith High Starch-hydrolytic Activity

Abstract

The fungal strain Mortierella alliacea YN-15 is an arachidonic acid producer that assimilates soluble starch despite having undetectable alpha-amylase activity. Here, a alpha-glucosidase responsible for the starch hydrolysis was purified from the culture broth through four-step column chromatography. Maltose and other oligosaccharides were less preferentially hydrolyzed and were used as a glucosyl donor for transglucosylation by the enzyme, demonstrating distinct substrate specificity as a fungal alpha-glucosidase. The purified enzyme consisted of two heterosubunits of 61 and 31 kDa that were not linked by a covalent bond but stably aggregated to each other even at a high salt concentration (0.5 M), and behaved like a single 92-kDa component in gel-filtration chromatography. The hydrolytic activity on maltose reached a maximum at 55 degrees C and in a pH range of 5.0-6.0, and in the presence of ethanol, the transglucosylation reaction to form ethyl-alpha-D-glucoside was optimal at pH 5.0 and a temperature range of 45-50 degrees C.

Purification and biochemical characterization of a novel α-glucosidase from Aspergillus niveus

“…It is noteworthy that gel filtration analysis of dimeric fungal a-glucosidases from Mortierella alliacea, Chaetomium thermophilum var. coprophilum and Aspergillus nidulans presented molecular weights of 92, 107 and 129 kDa (Tanaka et al 2002;Giannesi et al 2006;Kato et al 2002). Electrofocusing of the purified aglucosidase of A. niveus revealed a pI of 6.8, which is similar to value of the a-glucosidase from Mortierella alliceae (pI 6.3) (Tanaka et al 2002), in contrast, an a-glucosidase from Paecilomyces lilacinus presented a pI of 9.1 (Kobayashi et al 2003).…”

“…coprophilum and Aspergillus nidulans presented molecular weights of 92, 107 and 129 kDa (Tanaka et al 2002;Giannesi et al 2006;Kato et al 2002). Electrofocusing of the purified aglucosidase of A. niveus revealed a pI of 6.8, which is similar to value of the a-glucosidase from Mortierella alliceae (pI 6.3) (Tanaka et al 2002), in contrast, an a-glucosidase from Paecilomyces lilacinus presented a pI of 9.1 (Kobayashi et al 2003). The A. niveus a-glucosidase is a glycoprotein, and contains 29% carbohydrate, which is in agreement with other fungal a-glucosidases (Giannesi et al 2006;Yamamoto et al 2004).…”

“…4a, b). Most of the purified glucosidases from several other sources exhibit pH optima ranging from 4.5 to 7.0 (Frandsen and Svensson 1998;Yamasaki et al 1973;Oguma et al 1992;Giannesi et al 2006) and temperature optima ranging from 35 to 65°C (Kobayashi et al 2003;Tanaka et al 2002;Yamamoto et al 2004;Okuyama et al 2005). Thermal inactivation by incubating the enzyme in the absence of starch revealed that the purified enzyme was quite stable at 60°C (half-life 104 min), and decayed with a half-life of approximately 78 min at 65°C (Fig.…”

Purification and characterization of the α-glucosidase produced by thermophilic fungus Thermoascus aurantiacus CBMAI 756

“…2B, line 2), and suggesting that the enzyme is a trimeric protein. Oligomeric forms have been described for a number of microbial -glucosidases (Kato et al, 2002;Tanaka et al, 2002;Giannesi et al, 2006). Another possibility is that our protein have two subunits (112 and 39 kDa), which remain associated during the purifycation steps, and ~144 kDa (112+39) as an unprocessed polypeptide.…”

Purificação e caracterização bioquímica da α-glicosidase termoestável produzida por Thermoascus aurantiacus CBMAI 756

“…Várias Į-glicosidases purificadas de várias fontes exibem pH e temperatura ótimo ente 4,0 -7,5 e 50 -70°C, respectivamente (GINNESI et al, 2006;ANINDYAWATI et al, 1998;YAMAMOTO et al, 2004;BRAVO-TORRES et al, 2004;OKUYAMA et al, 2005;TANAKA et al, 2002;ZDZIEBLO;SYNOWIECKI, 2002;MARTINO et al, 2001;EZEJI;BAHL, 2006;IWATA et al, 2003).…”