ADENYLYLSULPHATE REDUCTASE FROM THE SULFATE-REDUCING ARCHAEON ARCHAEOGLOBUS-FULGIDUS - CLONING AND CHARACTERIZATION OF THE GENES AND COMPARISON OF THE ENZYME WITH OTHER IRON-SULFUR FLAVOPROTEINS

Adenylylsulphate (adenosine-5'-phosphosulphate, APS) reductase from th e extremely thermophilic sulphate-reducing archaeon Archaeoglobus fulg idus is an iron-sulphur flavoprotein containing one non-covalently bou nd flavin group, eight non-haem iron and six labile sulphide atoms per molecule. Reevaluation of the enzyme structure revealed the presence of two different subunits with molecular masses of 80 and 18.5 kDa. Th e subunits are arranged in an alpha(2) beta subunit structure. We have cloned and sequenced a 2.7 kb segment of DNA containing the genes for the alpha and beta subunits, which we designate aprA and aprB, respec tively. The two genes are separated by 17 bp and localized in the orde r aprBA. While a putative promoter could not be identified in the vici nity of aprBA a probable termination signal was found just downstream of the translation stop codon of aprA. The codon usage for aprBA shows strong preferences for G and C in the third codon position. aprA enco des a 73.3 kDa polypeptide, which shows significant overall similariti es with the flavoprotein subunits of the succinate dehydrogenases from Escherichia coli and Bacillus subtilis and the corresponding flavopro tein of E. coli fumarate reductase. Part of the homologous peptide str etches could be assigned to domains that are involved in the binding o f the substrate or of the FAD prosthetic group. aprB encodes a 17.1 kD a polypeptide representing an iron-sulphur protein, seven cysteine res idues of which are arranged in two clusters typical of ligands of the iron-sulphur centres in ([Fe3S4][Fe4S4]) 7-Fe ferredoxins.