Cloning, expression, and characterization of thermostable Manganese superoxide dismutase from Thermoascus aurantiacus var. levisporus
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-t...
Saved in:
Published in | The journal of microbiology Vol. 47; no. 1; pp. 123 - 130 |
---|---|
Main Authors | , , , |
Format | Journal Article |
Language | English |
Published |
Heidelberg
The Microbiological Society of Korea
01.02.2009
Springer Nature B.V 한국미생물학회 |
Subjects | |
Online Access | Get full text |
Cover
Loading…
Summary: | 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 μg/mg of protein and Fe was not detected in the purified enzyme. The enzyme was found to be inhibited by NaN
3
, but not by KCN or H
2
O
2
. 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 55°C and it exhibited maximal activity at pH 7.5. The enzyme was thermostable at 50 and 60°C and the half-life at 80°C was approximately 40 min. |
---|---|
Bibliography: | http://msk.jams.or.kr/jams/co/download/popup/poDownload.kci?storFileBean.orteFileId=FI0002204398 G704-000121.2009.47.1.011 |
ISSN: | 1225-8873 1976-3794 |
DOI: | 10.1007/s12275-008-0217-9 |