Putative ACP phosphodiesterase gene (acpD) encodes an azoreductase.
An FMN-dependent NADH-azoreductase of Escherichia coli was purified and analyzed for identification of the gene responsible for azo reduction by microorganisms. The N-terminal sequence of the azoreductase conformed to that of the acpD gene product, acyl carrier protein phosphodiesterase. Overexpression of the acpD gene provided the E. coli with a large amount of the 23-kDa protein and more than 800 times higher azoreductase activity. The purified gene product exhibited activity corresponding to that of the native azoreductase. The reaction followed a ping-pong mechanism requiring 2 mol of NADH to reduce 1 mol of methyl red (4'-dimethylaminoazobenzene-2-carboxylic acid) into 2-aminobenzoic acid and N,N'-dimethyl-p-phenylenediamine. On the other hand, the gene product could not convert holo-acyl carrier protein into the apo form under either in vitro or in vivo conditions. These data indicate that the acpD gene product is not acyl carrier protein phosphodiesterase but an azoreductase.
Nakanishi
M
M
Laboratory of Molecular Biochemistry, Department of Biomolecular Science, Faculty of Engineering, Gifu University, Gifu 501-1193, Japan.
Yatome
C
C
Ishida
N
N
Kitade
Y
Y
D000595:Amino Acid Sequence; D001483:Base Sequence; D002853:Chromatography, Liquid / Q000379:methods; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008969:Molecular Sequence Data; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D009601:Nitroreductases; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013057:Spectrum Analysis
D017931:DNA Primers; D009247:NADH, NADPH Oxidoreductases; D009601:Nitroreductases; C021106:azoreductase; D010727:Phosphoric Diester Hydrolases; C018754:(acyl-carrier-protein) phosphodiesterase