[Expression and characterization of the nonspecific azoreducase of Shewanella decolorationis S12].
OBJECTIVE
In order to identify the function of acpD in azoreduction, the gene was obtained from Shewanella decolorationis S12 and expressed in Escherichia coli TOP10 which showed little azoreduction activity.


METHODS
Gene cloning and expression of acpD were performed by pGM-T vector. Sequences were analyzed by DNAMAN software and azoreduction activity was assayed spectrophotometrically.


RESULTS
The deduced amino acid sequences from acpD of S. decolorationis S12 showed 61% identity to FMN-dependent NADH-azoreductase from E. coli JM109 with extremely conservative amino acid sequences at FMN binding sites. Moreover, the resulting recombinant E. coli TOP10 (pGMT-N) exhibited high azoreduction activity for azo dyes of different polarity by intact cells and their extracts. On the other hand, NADH was used as electron donor to drive azoreduction and FMN could enhance the reduction activity.


CONCLUSIONS
The product of acpD from S. decolorationis S12 is a nonspecific FMN-dependent NADH-azoreductase, which transfers electrons from NADH via FMN to azo bond, and exhibits high azoreduction activity under in vivo or in vitro conditions.
Chen
Xingjuan
X
South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, China. xingjuan_chen@yahoo.com.cn
Xu
Meiying
M
Li
Guangfei
G
Sun
Guoping
G
D000595:Amino Acid Sequence; D001391:Azo Compounds / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005486:Flavin Mononucleotide / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D008969:Molecular Sequence Data; D009243:NAD / Q000378:metabolism; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009601:Nitroreductases; D016415:Sequence Alignment; D020592:Shewanella / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity
D001391:Azo Compounds; D001426:Bacterial Proteins; D009243:NAD; D005486:Flavin Mononucleotide; D009247:NADH, NADPH Oxidoreductases; D009601:Nitroreductases; C021106:azoreductase