Purification and characterization of a thermostable glucoamylase from the thermophilic fungus Thermomyces lanuginosus

Abstract

Glucoamylase (1,4-alpha-D-glucan glucohydrolase, EC 3.2.1.3) was purified from the culture filtrates of the thermophilic fungus Thermomyces lanuginosus and was established to be homogeneous by a number of criteria. The enzyme was a glycoprotein with an average molecular weight of about 57 000 and a carbohydrate content of 10-12%. The enzyme hydrolysed successive glucose residues from the non-reducing ends of the starch molecule. It did not exhibit any glucosyltransferase activity. The enzyme appeared to hydrolyse maltotriose by the multi-chain mechanism. The enzyme was unable to hydrolyse 1,6-alpha-D-glucosidic linkages of isomaltose and dextran. It was optimally active at 70 degrees C. The enzyme exhibited increase in the Vmax. and decreased in Km values with increasing chain length of the substrate molecule. The enzyme was inhibited by the substrate analogue D-glucono-delta-lactone in a non-competitive manner. The enzyme inhibited remarkable resistance towards chemical and thermal denaturation.

Purification of extracellular amylolytic enzymes from the thermophilic fungus Thermomyces lanuginosus

“…an a-amylase, an a-glucosidase, and a glucoamylase. In static culture, Basaveswara Rao et al (1979) found that among four thermophilic fungi tested Thermomyces lanuginosus produced the highest specific activity of glucoamylase (52.1 U . mg protein-I), but in contrast to the present investigation the presence of other amylolytic activities was not reported.…”

“…Basaveswara Rao et al (1981) have purified a glucoamylase from the culture filtrate of Thermomyces lanuginosus. This enzyme was a glycoprotein with an average molecular weight of about 57 000 and a carbohydrate content of 10-12%.…”

“…This fungus has been suggested as an excellent test organism for the study of extracellular amylase from a thermophilic fungus (Barnett and Fergus 1971). Basaveswara Rao et al (1979) have identified a glucoamylase (EC 3.2.1.3) in the growth medium from cultivations on starch in both static cultures and shaken flask cultures. Taylor et al (1978) and Basaveswara Rao et al (198 1) have purified glucoamylases from submerged cultures and the former authors found traces of a-amylase activity.…”

CHARACTERIZATION OF [14C]-STARCH DEGRADATION BY RHIZOPUS GLUCOAMYLASE

“…Partial Purification of T. lanuginosus Glucoamylase. A culture of Thermomyces Zunuginosus ML-M was generously donated by V. Basaveswara Rao of the Indian Institute of Science, Bangalore, India, and grown in liquid culture as described (Basaveswara Rao et al 1981). Flasks were incubated at 50°C and shook at 250 rpm.…”

Effect of water activity on the thermal stability of Thermomyces lanuginosus xylanases for process time–temperature integration

“…Enzymes produced by thermophilic fungi are usually more thermostable than those produced by mesophilic fungi. Thermomyces lanuginosus is a thermophilic fungus that grows optimally between 48 and 52°C and secretes a variety of degradative thermozymes including: amylase (Odibo and Ulbrich-Hofmann, 2001), glucoamylase (Basaveswara Rao et al, 1981), hemicellulases (Lin et al, 1999;Singh et al, 2000a), lipase (Arima et al, 1972), phytase (Berka et al, 1998), serine-protease (Hasnain et al, 1992) and xylanases (Bennet et al, 1998;Gruber et al, 1998;Singh et al, 2000b;Singh et al, 2000c). Endo-b-1,4-xylanases (EC 3.2.1.8) from T. lanuginosus are extremely thermostable enzymes.…”