Interactions of [14C]phosphonoformic acid with renal cortical brush-border membranes. Relationship to the Na+-phosphate co-transporter.
Since phosphonoformic acid (PFA) acts as a specific competitive inhibitor of Na+-Pi co-transport across renal brush-border membrane (BBM), we employed the [14C]PFA as a probe to determine the mechanism of its interaction with rat renal BBM. The binding of [14C]PFA to BBM vesicles (BBMV), with Na+ present in extravesicular medium (Na+o), was time- and temperature-dependent. The replacement of Na+o with other monovalent cations reduced the PFA binding by -80%. Cl- was the most effective accompanying monovalent anion as NaCl for maximum PFA binding. The Na+o increased the apparent affinity of BBMV for [14C]PFA binding, but it did not change the maximum binding capacity. The maximum [14C]PFA binding was achieved at Na+o approximately equal to 50 mM. The extent of Na+-dependent [14C]PFA binding correlated (r = 0.98; p less than 0.01) with percent inhibition by an equimolar dose of PFA of the (Na+o greater than Na+i)-dependent BBMV uptake of 32Pi. Intravesicular Na+ (Na+i) decreased [14C]PFA binding, on BBMV, and this inhibition by Na+i was dependent on the presence of Na+o. The increase in Na+i, at constant [Na+]o, decreased the Vmax, but not the Km, for [14C]PFA binding on BBMV. Bound [14C]PFA was displaced from BBMV by phosphonocarboxylic acids proportionally (r = 0.99; p less than 0.05) to their ability to inhibit (Na+o greater than Na+i)-gradient-dependent Pi transport, whereas other monophosphonates, diphosphonates, L-proline, or D-glucose did not influence the [14C]PFA binding. The Na+-dependent binding of [14C]PFA and of [3H]phlorizin by BBMV was 10 times higher than binding of these ligands to renal basolateral membranes and to mitochondria. [14C]PFA probably binds onto the same locus on the luminal surface of BBM, where Pi and Na+ form a ternary complex with the Na+-Pi co-transporter.
D000818:Animals; D000838:Anions; D000998:Antiviral Agents / Q000378:metabolism; D002250:Carbon Radioisotopes; D002352:Carrier Proteins / Q000378:metabolism*; D002414:Cations, Monovalent; D017245:Foscarnet; D007672:Kidney Cortex / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008353:Mannitol / Q000494:pharmacology; D008871:Microvilli / Q000378:metabolism*; D009943:Organophosphorus Compounds / Q000378:metabolism*; D010746:Phosphonoacetic Acid / Q000031:analogs & derivatives / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D011919:Rats, Inbred Strains; D050608:Sodium-Phosphate Cotransporter Proteins; D027981:Symporters
D000838:Anions; D000998:Antiviral Agents; D002250:Carbon Radioisotopes; D002352:Carrier Proteins; D002414:Cations, Monovalent; D009943:Organophosphorus Compounds; D050608:Sodium-Phosphate Cotransporter Proteins; D027981:Symporters; D017245:Foscarnet; D008353:Mannitol; D010746:Phosphonoacetic Acid