To maintain acid-base homeostasis the kidney excretes the excess acid produced from metabolism by secreting H plus from the tubular cell into the lumen. Because of the complexity of the renal architecture the cellular processes that underlie H plus secretion were studied in a model urinary epithelium, the turtle bladder. There we (and others) found that H plus secretion is regulated by CO2, carbonic anhydrase activity, aldosterone and the transepithelial electrochemical gradient. We recently found that the H plus pump which is located in the luminal border is a reversible proton-translocating ATPase. This background information is sufficiently detailed to allow us to start a study of the subcellular and molecular details of the H plus pump. To this end we prepared membrane fractions enriched in luminal and basolateral fragments of the cell membrane of turtle bladder. We plan to study H plus transport in these vesicles under well-defined conditions. The relation between H plus transport and ATP hydrolysis will be studied quantitatively. We have also started to isolate the H plus pump from the outer medulla of the kidney. Using these two sources we plan to purify the H plus pump and examine its molecular details using reconstitution as an assay for functional units. These studies should help to lay a molecular foundation for our understanding of urinary acidification in health and in the various disease states in which it is deranged such as renal failure hyperaldosteronism and tubular disease.
