Recombinant Ca2+–ATPase was expressed in Saccharomyces cerevisiae with a biotin-acceptor domain linked to its C–terminus by a thrombin cleavage site. We obtained 200 μg of ~ 70% pure recombinant sarcoendoplasmic reticulum Ca2+–ATPase isoform 1a (SERCA1a) from a 6–L yeast culture. The catalytic cycle of SERCA1a was followed in real time using rapid scan FTIR spectroscopy. Different intermediate states (Ca2E1P and Ca2E2P) of the recombinant protein were accumulated using different buffer compositions. The difference spectra of their formation from Ca2E1 had the same spectral features as those from the native rabbit SERCA1a. The enzyme-specific activity for the active enzyme fraction in both samples was also similar. The results show that the recombinant protein obtained from the yeast-based expression system has similar structural and dynamic properties as native rabbit SERCA1a. It is now possible to apply this expression system together with IR spectroscopy to the investigation of the role of individual amino acids.