Primary structure of xylene monooxygenase: similarities to and differences from the alkane hydroxylation system

Abstract

Xylene monooxygenase, encoded by the TOL plasmid of Pseudomonas putida, catalyzes the oxidation of toluene and xylenes and consists of two different subunits encoded by xylA and xylM. In this study, the complete nucleotide sequences of these genes were determined and the amino acid sequences of the xylA and xylM products were deduced. The XylM sequence had a 25% homology with alkane hydroxylase, which catalyzes the omega-hydroxylation of fatty acids and the terminal hydroxylation of alkanes. The sequence of the first 90 amino acids of XylA exhibited a strong similarity to the sequence of chloroplast-type ferredoxins, whereas the rest of the XylA sequence resembled that of ferredoxin-NADP+ reductases. Based on this information, the structure and function of xylene monooxygenase were deduced. XylM may be a catalytic component for the hydroxylation of the carbon side chain of toluene and xylenes and, as is the alkane hydroxylase protein, may be a membrane-bound protein containing ferrous ion as a prosthetic group. XylA may have two domains consisting of an N-terminal region similar to chloroplast-type ferredoxins and a C-terminal region similar to ferredoxin-NADP+ reductases. The ferredoxin portion of XylA may contain a [2Fe-2S] cluster and reduce the oxidized form of the XylM hydroxylase. The activity determined by the C-terminal region of the XylA sequence may be the reduction of the oxidized form of ferredoxin by concomitant oxidation of NADH.