Biochemical and genetic characterization of 2-carboxybenzaldehyde dehydrogenase, an enzyme involved in phenanthrene degradation by Nocardioides sp. strain KP7

Author:

Iwabuchi T1,Harayama S1

Affiliation:

1. Marine Biotechnology Institute, Kamaishi Laboratories, Iwate, Japan.

Abstract

2-Carboxybenzaldehyde dehydrogenase from the phenanthrene-degrading bacterium Nocardioides sp. strain KP7 was purified and characterized. The purified enzyme had a molecular mass of 53 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 205 kDa by gel filtration chromatography. Thus, the homotetramer of the 53-kDa subunit constituted an active enzyme. The apparent Km and kcat values of this enzyme for 2-carboxybenzaldehyde were 100 microM and 39 s(-1), respectively, and those for NAD+ were 83 microM and 32 s(-1), respectively. The structural gene for this enzyme was cloned and sequenced. The length of the gene was 1,455 bp. The nucleotide sequence of the 10,279 bp of DNA around the gene for 2-carboxybenzaldehyde dehydrogenase was also determined, and seven open reading frames were found in this DNA region. These were the genes for 1-hydroxy-2-naphthoate dioxygenase (phdI) and trans-2'-carboxybenzalpyruvate aldolase (phdJ), orf1, the gene for 2-carboxybenzaldehyde dehydrogenase (phdK), orf2/orf3, and orf4. The amino acid sequence of the orf1 product was similar to that of the aromatic hydrocarbon transporter gene (pcaK) in Pseudomonas putida PRS2000. The amino acid sequence of the orf4 product revealed a similarity to cytochrome P-450 proteins. The region between phdK and orf4 encoded orf2 and orf3 on different strands. The amino acid sequences of the orf2 and orf3 products exhibited no significant similarity to the reported sequences in protein databases.

Publisher

American Society for Microbiology

Subject

Molecular Biology,Microbiology

Reference36 articles.

1. Variations and coding features of the sequence spanning the replication terminus of Bacillus subtilis 168 and W23 chromosomes;Ahn K. S.;Gene,1991

2. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding;Bradford M. M.;Anal. Biochem.,1976

3. Superpolylinkers in cloning and expression vectors;Brosius J.;DNA,1989

4. Cerniglia C. E. and M. A. Heitkamp. 1989. Microbial degradation of polycyclic aromatic hydrocarbons (PAH) in the aquatic environment p. 41-68. In U. Varanasi (ed.) Metabolism of polycyclic aromatic hydrocarbons in the aquatic environment. CRC Press Inc. Boca Raton Fla.

5. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid;Chang A. C. Y.;J. Bacteriol.,1978

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