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Characterization of a benzoylformate decarboxylase and a NAD+/NADP+‐dependent benzaldehyde dehydrogenase from Pseudomonas stutzeri ST‐201
Author(s) -
Saehuan Choedchai,
Rojanarata Theerasak,
Wiyakrutta Suthep,
Meevootisom Vithaya,
McLeish Michael J.
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.1008.4
Subject(s) - pseudomonas putida , pseudomonas stutzeri , nad+ kinase , dehydrogenase , biochemistry , chemistry , benzaldehyde , enzyme , operon , stereochemistry , biology , bacteria , gene , mutant , catalysis , genetics
Both D‐phenylglycine aminotransferase and benzoylformate decarboxylase (BFDC) activity were detected in cultures of P. stutzeri ST‐201 following induction with D‐phenylglycine. Conversely, induction with benzoylformate enhanced only BFDC activity. Colony hybridization was used to identify two genomic DNA fragments which, when combined, contained two open reading frames whose gene products were characterized as a benzoylformate decarboxylase and a NAD + /NADP + ‐dependent benzaldehyde dehydrogenase. The kinetic properties of the BFDC was similar to those of the BFDC found in the mandelate pathway of P. putida ATCC 12633. However, the K m values for NAD(P) + of the benzaldehyde dehydrogenase were significantly higher than those of the P. putida enzyme. Based on sequence and structural alignments this was likely to be due to a change from leucine to arginine at position 175. Given that P. stutzeri ST‐201 was unable to grow on either isomer of mandelate, and that sequencing indicated that dpgB did not form part of an operon , it appears that the two enzymes form part of a D‐phenylglycine, rather than mandelate, degrading pathway. This work is supported in part by the CHED and Faculty of Graduate Studies, Mahidol University, and granted by TRF through the RGJ‐Ph.D. Program No. Ph.D/0023/2548. The work was also supported by NSF EF 0425719.