Open AccessBiochemical characterization of α‐amylase from the yeast Cryptococcus flavus
Author(s) -
Wanderley Kenya J,
Torres Fernando A.G,
Moraes Lı́dia M.P,
Ulhoa Cirano J
Publication year - 2004
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1016/s0378-1097(03)00955-8
Subject(s) - amylopectin , maltose , pullulan , amylase , maltotriose , starch , biochemistry , size exclusion chromatography , chemistry , amylose , enzyme assay , chromatography , molecular mass , polyacrylamide gel electrophoresis , enzyme , yeast , yeast extract , aspergillus flavus , polysaccharide , food science , fermentation
During our screening of amylolytic microorganisms from Brazilian fruits, we isolated a yeast strain classified as Cryptococcus flavus . When grown on starch‐containing medium this strain exhibited the highest amylase production after 24 h of cultivation. The extracellular amylase from C. flavus was purified from the culture broth by a single step using chromatography on a Sephacryl S‐100 column. The enzyme was purified 16.14‐fold with a yield of 50.21% of the total activity. The purified enzyme was a glycoprotein with an apparent molecular mass of 75 and 84.5 kDa as estimated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and gel filtration, respectively. The enzyme lost approximately 50% of the molecular mass after treatment with glycosidases. The major end products of starch, amylose, amylopectin, pullulan and glycogen were maltose and maltotriose. The K m value for the pure enzyme was 0.056 mg ml −1 with soluble starch as the substrate. Enzyme activity was optimal at pH 5.5 and 50°C. The enzyme retained 90% of the activity after incubation at 50°C for 60 min and was inhibited by Cu 2+ , Fe 2+ and Hg 2+ .