Purification and Properties of p-Hydroxybenzoate Hydroxylases from Rhodococcus Strains

A. P. Jadan^1,2, W. J. H. van Berkel², L. A. Golovleva^1*, and E. L. Golovlev¹

¹Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, Pushchino, Moscow Region, Russia; fax: (095) 923-3602; E-mail: Golovleva@ibpm.serpukhov.su

²Laboratory of Biochemistry, Department of Agrotechnology and Food Sciences, Wageningen University, Dreijenlaan 3, 6703 HA Wageningen, The Netherlands

^* To whom correspondence should be addressed.

Received December 28, 2000; Revision received February 5, 2001

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Gram-positive bacteria of the genus Rhodococcus catabolize p-hydroxybenzoate (PHB) through the initial formation of 3,4-dihydroxybenzoate. High levels of p-hydroxybenzoate hydroxylase (PHBH) activity are induced in six different Rhodococcus species when these strains are grown on PHB as sole carbon source. The PHBH enzymes were purified to apparent homogeneity and appeared to be homodimers of about 95 kD with each subunit containing a relatively weakly bound FAD. In contrast to their counterparts from gram-negative microorganisms, the Rhodococcus PHBH enzymes prefer NADH to NADPH as external electron donor. All purified enzymes were inhibited by Cl^- and for five of six enzymes more pronounced substrate inhibition was observed in the presence of chloride ions.

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KEY WORDS: Rhodococcus species, flavoprotein, p-hydroxybenzoate hydroxylase, substrate inhibition

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