Calcium pump, high-affinity Ca²⁺-ATPase, and other ATPases in dog antrum smooth muscle plasma membrane

The plasma membrane-enriched fraction from dog antrum smooth muscle is enriched in ATP-dependent azide-insensitive Ca2+ uptake (0.3-0.4 μM Ca2+ required for half-maximal activity), a high-affinity Ca2+-ATPase (K(m) of 0.3-0.8 μM for Ca2+), a low-affinity Ca2+-ATPase (K(m) for 250-400 μM for Ca2+) and a Mg2+-ATPase. Studies using membranes washed with EDTA and assay media treated with Chelex 100 showed that the high-affinity Ca2+-ATPase did not depend on contaminating Mg2+. Thus, whereas the ATP-dependent Ca2+ uptake had an absolute requirement for Mg2+, the Ca2+-ATPases did not. Studies using γ-irradiation showed that the protein responsible for the ATP-dependent Ca2+ uptake was inactivated at significantly lower doses of radiation than the three ATPases. The Ca2+ uptake and the high-affinity Ca2+-ATPase also differed in their inhibition by calmodulin antagonists and 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid. Thus it is unlikely that the high-affinity Ca2+ ATPase by itself is responsible for the ATP-dependent Ca2+ uptake.