CHARACTERIZATION OF THE ADENOSINE-TRIPHOSPHATASE ACTIVITY OF THE PERIPLASMIC HISTIDINE PERMEASE, A TRAFFIC ATPASE (ABC TRANSPORTER)

The superfamily of traffic ATPases (ABC transporters) includes bacteri al periplasmic transport systems (permeases) and eukaryotic transporte rs. The histidine permease of Salmonella typhimurium is composed of a membrane-bound complex (HisQMP(2)) containing four subunits, and of a soluble receptor, the histidine-binding protein (HisJ). Transport is e nergized by ATP, In this article the ATPase activity of HisQMP(2) has been characterized, using a novel assay that is independent of transpo rt. The assay uses Mg2+ ions to permeabilize membrane vesicles or prot eoliposomes, thus allowing access of ATP to both sides of the bilayer, HisQMP(2) displays a low level of intrinsic ATPase activity in the ab sence of HisJ; unliganded HisJ stimulates the activity and liganded Hi sJ stimulates to an even higher level, All three levels of activity di splay positive cooperativity for ATP with a Hill coefficient of 2 and a K-0.5 value of 0.6 mM. The activity has been characterized with resp ect to pH, salt, phospholipids, substrate, and inhibitor specificity, Free histidine has no effect. The activity is inhibited by orthovanada te, but not by N-ethylmaleimide, bafilomycin A(1), or ouabain. Several nucleotide analogs, ADP, 5'-adenylyl-beta,gamma-imidodiphosphate, ade nosine 5'-(beta,gamma-imino)triphosphate, and adenosine 5'-O-(3-thio)t riphosphate, inhibit the activity. Unliganded HisJ does not compete wi th liganded HisJ for the stimulation of the ATPase activity of HisQMP( 2).