CLONING OF THE 3'-PHOSPHOADENYLYL SULFATE REDUCTASE AND SULFITE REDUCTASE GENES FROM ESCHERICHIA-COLI K-12

The structural genes for 3'-phosphoadenylyl sulfate (PAPS) reductase (cysH) and sulfite reductase (a and β subunits; EC 1.8.1.2) (cysI and cysJ) of Escherichia coli K-12 have been cloned by complementation. pCYSI contains two PstI fragments (18.3 and 2.9kb) which complement cysH−, cysI−, and cysJ− mutants. Subcloning showed that the cysH gene is located on a 1.6-kb ClaI subfragment (pCYSI-3) whereas cysI and most of cysJ are carried on a 3.7-kb ClaI subfragment (pCYSI-5). The PAPS reductase gene is closely linked to the sulfite reductase genes, but its expression is regulated by a unique promoter. The cysI and cysJ genes, on the other hand, are transcribed as an operon and the promoter precedes the cysI gene. Maxicell analysis demonstrated that pCYSI encodes three polypeptides of Mr 27000, 57000, and 60000, in addition to the tetracycline-resistance determinant. The 60- and 57-kDa proteins are most likely the α and β subunits, respectively, of E. coli sulfite reductase while the 27-kDa protein is putatively identified as PAPS reductase. Preliminary data suggest that the α and β subunits of sulfite reductase are encoded by cysI and cysJ, respectively.