A functional 4-hydroxysalicylate/hydroxyquinol degradative pathway gene cluster is linked to the initial dibenzo-p-dioxin pathway genes in Sphingomonas sp strain RW1

The bacterium Sphingomonas sp. strain RW1 is able to use dibenzo-p-dioxin, dibenzofuran, and several hydroxylated derivatives as sole sources of carbo n and energy. We have determined and analyzed the nucleic acid sequence of a 9,997-bp HindIII fragment downstream of cistrons dxnA1A2, which encode th e dioxygenase component of the initial dioxygenase system of the correspond ing catabolic pathways. This fragment contains 10 colinear open reading fra mes (ORFs), apparently organized in one compact operon. The enzymatic activ ities of some proteins encoded by these genes were analyzed in the strain R W1 and, after hyperexpression, in Escherichia coil, The first three ORFs of the locus, designated dxnC, ORF2, and fdx3, specify a protein with a low h omology to bacterial siderophore receptors, a polypeptide representing no s ignificant homology to known proteins, and a putative ferredoxin, respectiv ely. dxnD encodes a 69-kDa phenol monooxygenase like protein with activity for the turnover of 4-hydroxysalicylate, and dxnE codes for a 37-kDa protei n whose sequence and activity are similar to those of known maleylacetate r eductases. The following gene, dxnF, encodes a 33-KDa intradiol dioxygenase which efficiently cleaves hydroxyquinol, yielding maleylacetate, the ketof orm of 3-hydroxy-cis,cis-muconate. The heteromeric protein encoded by dxnGH is a 3-oxoadipate succinyl coenzyme A (succinyl-CoA) transferase, whereas dxnI specifies a protein exhibiting marked homology to acetyl-CoA acetyltra nsferases (thiolases), The last ORF of the sequenced fragment codes for a p utative transposase. DxnD, DxnF, DxnE, DxnGH, and DxnI (the activities of m ost of them have also been detected in strain RW1) thus form a complete 4-h ydroxysalicylate/hydroxyquinol degradative pathway. A route for the mineral ization of the growth substrates 3-hydroxydibenzofuran and 2-hydroxydibenzo -p-dioxin in Sphingomonas sp, strain RW1 thus suggests itself.