Open Access

Andreas Barth

• Time-resolved Fourier transform infrared difference spectra of the phosphoenzyme conversion and Ca(2+) release reaction (Ca(2)E(1)-P --> E(2)-P) of the sarcoplasmic reticulum Ca(2+)-ATPase were recorded at pH 7 and 1 degrees C in H(2)O and (2)H(2)O. In the amide I spectral region, the spectra indicate backbone conformational changes preserving conformational changes of the preceding phosphorylation step. beta-sheet or turn structures (band at 1685 cm(-1)) and alpha-helical structures (band at 1653 cm(-1)) seem to be involved. Spectra of the model compound EDTA for Ca(2+) chelation indicate the assignment of bands at 1570, 1554, 1411 and 1399 cm(-1) to Ca(2+) chelating Asp and Glu carboxylate groups partially shielded from the aqueous environment. In addition, an E(2)-P band at 1638 cm(-1) has been tentatively assigned to a carboxylate group in a special environment. A Tyr residue seems to be involved in the reaction (band at 1517 cm(-1) in H(2)O and 1515 cm(-1) in (2)H(2)O). A band at 1192 cm(-1) was shown by isotopic replacement in the gamma-phosphate of ATP to originate from the E(2)-P phosphate group. This is a clear indication that the immediate environment of the phosphoenzyme phosphate group changes in the conversion reaction, altering phosphate geometry and/or electron distribution.