DISSIMILATORY SULFITE REDUCTASE FROM ARCHAEOGLOBUS-FULGIDUS - PHYSICOCHEMICAL PROPERTIES OF THE ENZYME AND CLONING, SEQUENCING AND ANALYSISOF THE REDUCTASE GENES

A dissimilatory sulphite reductase was isolated from the extremely the rmophilic dissimilatory sulphate-reducing archaeon Archaeoglobus fulgi dus. In common with other dissimilatory sulphite reductases thus far c haracterized, the enzyme has an alpha2beta2-structure and contains sir ohaem, non-haem iron atoms and acid labile sulphide. The oxidized enzy me exhibited absorption maxima at 281, 394, 545 and 593 nm with a weak band around 715 nm. We have cloned and sequenced the genes for the al pha and beta subunits of this enzyme, which we designate dsrA and dsrB , respectively. They are contiguous in the order dsrA dsrB and probabl y comprise an operon, since dsrA is preceded by sequences characterist ic of promoters in methanogenic archaea, and dsrB is followed by a seq uence resembling termination signals in extremely thermophilic sulphur -dependent archaea. dsrA and dsrB encode 47-4 kDa and 41.7 kDa peptide s, which have 25.6 % amino acid sequence identity, indicating that the y may have arisen by duplication of an ancestral gene. Each deduced pe ptide contains cysteine clusters resembling those postulated to bind s irohaem-[Fe4S4] complexes in sulphite reductases and nitrite reductase s from other species. The dsrB encoded peptide lacks a single cysteine residue in one of the two clusters, suggesting that only the alpha su bunit binds a sirohaem-[Fe4S4] complex, and chemical analyses showed t he presence of only two sirohaems per alpha2beta2 enzyme molecule. Bot h deduced peptides also contain an arrangement of cysteine residues ch aracteristic of [Fe4S4] ferredoxins, and chemical analyses were consis tent with the presence of six [Fe4S4] clusters per alpha2beta2 enzyme molecule, two of which would be expected to be associated with sirohae m while the other four could bind to the ferredoxin-like sites.