Journal of Microbiology

Research Support, Non-U.S. Gov'ts

Cloning, Expression, and Characterization of Thermostable Manganese Superoxide Dismutase from Thermoascus aurantiacus var. levisporus: Ning-Ning Song , Yan Zheng , Shi-Jin E , Duo-Chuan Li; J. Microbiol. 2009;47(1):123-130. Published online February 20, 2009; DOI: https://doi.org/10.1007/s12275-008-0217-9

Abstract: A superoxide dismutase (SOD) gene of Thermoascus aurantiacus var. levisporus, a thermophilic fungus, was cloned, sequenced, and expressed in Pichia pastoris and its gene product was characterized. The coding sequence predicted a 231 residues protein with a unique 35 amino acids extension at the N-terminus indicating a mitochondrial-targeting sequence. The content of Mn was 2.46 ug/mg of protein and Fe was not detected in the purified enzyme. The enzyme was found to be inhibited by NaN3, but not by KCN or H2O2. These results suggested that the SOD in Thermoascus aurantiacus var. levisporus was the manganese superoxide dismutase type. In comparison with other MnSODs, all manganese-binding sites were also conserved in the sequence (H88, H136, D222, H226). The molecular mass of a single band of the enzyme was estimated to be 21.7 kDa. The protein was expressed in tetramer form with molecular weight of 68.0 kDa. The activity of purified protein was 2,324 U/mg. The optimum temperature of the enzyme was 55oC and it exhibited maximal activity at pH 7.5. The enzyme was thermostable at 50 and 60oC and the half-life at 80oC was approximately 40 min.

A Specific Short Dextrin-Hydrolyzing Extracellular Glucosidase from the Thermophilic Fungus Thermoascus aurantiacus 179-5: Ana Flavia Azevedo Carvalho , Aline Zorzetto Gonclves , Roberto da Silva , Eleni Gomes; J. Microbiol. 2006;44(3):276-283.; DOI: https://doi.org/2385 [pii]

Abstract: The thermophilic fungus Thermoascus aurantiacus 179-5 produced large quantities of a glucosidase which preferentially hydrolyzed maltose over starch. Enzyme production was high in submerged fermentation, with a maximal activity of 30 U/ml after 336 h of fermentation. In solid-state fermentation, the activity of the enzyme was 22 U/ml at 144 h in medium containing wheat bran and 5.8 U/ml at 48 h when cassava pulp was used as the culture medium. The enzyme was specific for maltose, very slowly hydrolyzed starch, dextrins (2-7G) and the synthetic substrate (α-PNPG), and did not hydrolyze sucrose. These properties suggest that the enzyme is a type II α-glucosidase. The optimum temperature of the enzyme was 70?. In addition, the enzyme was highly thermostable (100% stability for 10 h at 60? and a half-life of 15 min at 80?), and stable within a wide pH range.

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