THE ORF1 OF THE GENTAMICIN-RESISTANCE OPERON (AAC) OF PSEUDOMONAS-AERUGINOSA ENCODES ADENOSINE 5'-PHOSPHOSULFATE KINASE

The gentamicin-resistance operon of Pseudomonas aeruginosa (aac) conta ins two cistrons for which only the second gene product has an identif ied function. The 813bp second cistron (ORF2) encodes a protein that c onfers gentamicin resistance by catalysis of the transfer of an acetyl group from acetyl Coenzyme A to gentamicin. The first open reading fr ame (ORF1) encodes a 23.9 kDa protein that we have found, by enzyme ac tivity and immunological reactivity, to be adenosine-5'-phosphosulphat e (APS) kinase. APS kinase catalyses the transfer of the gamma phospho ryl group of ATP to the 3'-hydroxyl group of APS. The 70% sequence sim ilarity between the Pseudomonas and Escherichia coli APS kinases sugge sts that the Pseudomonas enzyme may catalyse phosphoryl transfer to th e 3'-hydroxyl group of other nucleotides such as dephosphocoenzyme A, as does the purified E. coli APS kinase. In extracts of pseudomonad ce lls we have also detected a higher molecular mass (70 kDa) protein tha t cross-reacts with an anti-E. coli APS kinase antibody. This cross-re active protein is also present in Pseudomonas strains lacking the gent amicin-resistance plasmid, and apparently reflects an APS kinase analo gous to the nodQ-encoded high-molecular-weight APS kinase present in R hizobium meliloti. Production of the Pseudomonas aac APS kinase was re pressed by cysteine when expressed in E. coli, as is E. coli APS kinas e. However, cysteine did not repress production of the Pseudomonas enz yme when the aac ORF1-encoded enzyme was expressed in a Pseudomonas st rain, indicating differential regulation of gene expression in the two organisms.