ePrints@IIScePrints@IISc Home | About | Browse | Latest Additions | Advanced Search | Contact | Help

Purification and properties of benzoate-4-hydroxylase from a soil pseudomonad

Reddy, C Channa and Vaidyanathan, CS (1971) Purification and properties of benzoate-4-hydroxylase from a soil pseudomonad. In: Archives of Biochemistry and Biophysics, 177 (2). pp. 488-498.

[img] PDF
fultext.pdf - Published Version
Restricted to Registered users only

Download (913kB) | Request a copy
Official URL: http://www.sciencedirect.com/science?_ob=ArticleUR...


Benzoate-4-hydroxylase from a soil pseudomonad was isolated and purified about 50-fold. Polyacrylamide gel electrophoresis of this enzyme preparation showed one major band and one minor band. The approximate molecular weight of the enzyme was found to be 120,000. Benzoate-4-hydroxylase was most active around pH 7.2. The enzyme showed requirements for tetrahydropteridine as the cofactor and molecular oxygen as the electron acceptor. NADPH, NADH, dithiothreitol, β-mercaptoethanol, and ascorbic acid when added alone to the reaction mixture did not support the hydroxylation reaction to any significant extent. However, when these compounds were added together with tetrahydropteridine, they stimulated the hydroxylation. This stimulation is probably due to the reduction of the oxidized pteridine back to the reduced form. This enzyme was activated by Fe2+ and benzoate. It was observed that benzoate-4-hydroxylase could catalyze the oxidation of NADPH in the presence of benzoate,p-aminobenzoate, p-nitrobenzoate, p-chlorobenzoate, and p-methylbenzoate, with only benzoate showing maximum hydroxylation. Inhibition studies with substrate analogs and their kinetic analysis revealed that the carboxyl group is involved in binding the substrate to the enzyme at the active center. The enzyme catalyzed the conversion of 1 mol of benzoate to 1 mol of p-hydroxybenzoate with the consumption of slightly more than 1 mol of NADPH and oxygen.

Item Type: Journal Article
Publication: Archives of Biochemistry and Biophysics
Publisher: Elsevier Science
Additional Information: Copyright of this article belongs to Elsevier Science.
Department/Centre: Division of Biological Sciences > Biochemistry
Date Deposited: 11 Dec 2009 11:37
Last Modified: 19 Sep 2010 05:47
URI: http://eprints.iisc.ac.in/id/eprint/23860

Actions (login required)

View Item View Item