Catabolism of acenaphthylene by Rhizobium sp. CU-A1

Abstract

Rhizobium sp. CU-A1 was previously shown to mineralize acenaphthylene via the formation of acenaphthenequinone, naphthalene-1,8-dicarboxylic acid and gentisic acid as the intermediates. In this work, several additional intermediates were isolated from the extracts of the culture broth of its blocked mutants, purified by thin-layer, silica gel column or high-performance liquid chromatography and identified by gas chromatography-mass spectrometry. As a result, a complete pathway for the catabolism of acenaphthylene by Rhizobium sp. CU-A1 is proposed as follow: the initial reaction is the incorporation of two oxygen atoms into acenaphthylene molecule to from acenaphthenediol which then further oxidized to acenaphthenequinone. Subsequent dioxygenation would yield a ring fission product, naphthalene-1,8-dicarboxylic acid. A possible decarboxylation would give a novel intrmediate, 1-naphthoic acid, which would further be oxidized to 1,2-dihydroxynaphthalene, salicylic acid and gentisic acid, respectively. Then, hydroxylation of gentisic acid, following by ring cleavage, would give central intermediates in the TCAcycle via maleylpyruvate or fumarylpyruvate formation. Crude cell free extract prepared from this organism grown in mineral medium supplemented with protocatechuic acid and induced by acenaphthylene showed ability to oxidize acenaphthylene to the corresponding oxidation products. The enzyme catalyzing the initial reaction of acenaphthylene degradation, acenaphthylene dioxygenase, was separated into three components, designated as components A, B, and C by gel filtration chromatography. All three components were essentially required for the activity giving acenaphthenediol as a product. Component A, terminal oxygenase, was a dimer with two subunits with molecular mass of 45 and 22 kDa. Component B, reductase, and C, ferridoxin, consisted of a single polypeptide with a molecular mass of 48.2 and 9.8 kDa, respectively. Component A and C had UV/visible spectra of Rieske type iron-sulfur center while component B had a UV/vivible spectrum of flavoprotein. Acenaphthylene dioxygenase from Rhizobium sp. CU-A1 was presumed to be a class IIB dioxygenase which had ring hydroxylating and ring fission activity.