Deduced amino acid sequence and E1–E2 equilibrium of the sarcoplasmic reticulum Ca2+‐ATPase of frog skeletal muscle Comparison with the Ca2+‐ATPase of rabbit fast twitch muscle

Abstract

The cDNA encoding a Ca²⁺-transport ATPase of frog (Rana esculenia) skeletal muscle was isolated and characterized. The deduced amino acid sequence, consisting of 994 residues, showed 89% identity to the fast twitch muscle sarcoplasmic reticulum Ca²⁺-ATPases of chicken and rabbit. Northern blot analysis using a fragment of this cDNA as probe detected a 5.0 kb message in frog skeletal muscle but did not detect any mRNA encoding sarcoplasmic reticulum Ca²⁺-ATPase in frog cardiac muscle. The enzymatic properties of the amphibian skeletal muscle Ca²⁺-ATPase were compared with those of the rabbit fast twitch muscle Ca²⁺-ATPase by functional expression of the cDNAs in COS-1 cells. The amphibian Ca²⁺-ATPase displayed a reduced apparent affinity for Ca²⁺ and an increased apparent affinity for the inhibitors, vanadate and thapsigargin, relative to the mammalian enzyme. This may be explained by a mechanism in which relatively more of the E₂ conformation accumulated ¹n the frog Ca²⁺-ATPase than in the mammalian enzyme.