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中国生物工程杂志

China Biotechnology
China Biotechnology  2010, Vol. 30 Issue (11): 24-29    DOI:
    
Prokaryotic Expression and Functional Analysis of Maize BAS1(2-Cys peroxiredoxinA)
CHENG Lei, WANG Lei-lei, CHENG Bei-jiu, FAN Jun
The Provincial Key Lab of Crop Science, Anhui Agricultural University, School of Life Science, Anhui Agricultural University, Hefei 230036, China
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Abstract  

The plant 2-Cys peroxiredoxin BAS1 is a thiol-dependent peroxidase capable of reducing hydrogen peroxide through the reactive catalytic cysteine, which is regenerated by NADPH dependent thioredoxin reductase from chloroplast (NTRC). Maize contains two BAS1, 2-Cys peroxiredoxin A (2-Cys PrxA) and 2-Cys peroxiredoxin B (2-Cys PrxB).The BAS1 gene encoding the mature 2-Cys PrxA from maize leaves was amplified by RT-PCR, and Cys34 was mutated into Ser34 in the BAS1 by site-directed mutagenesis. The SDS-PAGE analysis displayed that the subunit of the purified BAS1 wild type and mutant C34S has the apparent molecular mass about 23kDa. The protein binding assay showed the purified BAS1 mutant C34S is associated with the purified NTRC through the intermolecular disulfide bridge. Non-reducing SDS-PAGE analysis proved the purified BAS1 wild type formed the dimer through the intermolecular disulfide bridge, whereas the purified BAS1 mutant C34S formed the monomer. The results of modification BAS1 with thiol-reactive labeling reagent AMS and the activity analysis identified that the reducing BAS1 is essential for the catalysis of the reduction of hydrogen peroxide, which is kept by NTRC and its cofactor NADPH.



Key wordsBAS1(2-Cys/peroxiredoxinA)      NADPH thioredoxin reductase from chloroplast(NTRC)      Maize (Zea mays)      Functional analysis     
Received: 19 August 2010      Published: 19 November 2010
ZTFLH:  Q554+.6  
Cite this article:

CHENG Lei, WANG Lei-lei, CHENG Bei-jiu, FAN Jun. Prokaryotic Expression and Functional Analysis of Maize BAS1(2-Cys peroxiredoxinA). China Biotechnology, 2010, 30(11): 24-29.

URL:

https://manu60.magtech.com.cn/biotech/     OR     https://manu60.magtech.com.cn/biotech/Y2010/V30/I11/24

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