This article summarizes physiological and molecular studies of the colonic H+, K+-ATPase and active potassium (K) absorption in the rat distal colon. Active K absorption is energized by an apical membrane H+, K+-ATPase that is a member of the gene family of P-type ATPases. Physiological experiments of active K absorption and enzymatic analysis of H+, K+-ATPase provide compelling evidence for 2 distinct H+, K+-ATPases with a spatial distribution to surface and crypt epithelial cells: an ouabain-sensitive isoform is present in crypt cells and an ouabain-insensitive one is present in surface cells. An alpha subunit (HKcalpha1) has been cloned from the rat colon and its message and protein are selectively located in surface epithelial cells and apical membrane of surface epithelial cells, respectively. A beta subunit (HKcbeta) has also been cloned from the rat colon and may represent the beta subunit for the colonic H+, K+-ATPase. Expression studies have yielded conflicting data: studies with HKcalpha1 and HKcbeta cDNAs that have assessed H+, K+-ATPase activity have concluded that these cDNAs encode the ouabain-insensitive isoform. In contrast, studies with HKcalpha1 and other beta subunits that have expressed 86Rb uptake have yielded evidence for a ouabain-sensitive isoform. Although both aldosterone and dietary K depletion stimulate active K absorption, only the former involves the regulation of HKcalpha1 at a posttranscriptual level.