Different classes of tryptophan residues involved in the conformational changes characteristic of the sarcoplasmic reticulum Ca2(+)-ATPase cycle.
Various classes of tryptophan residues in the Ca2(+)-ATPase of sarcoplasmic reticulum membranes have been distinguished on the basis of their sensitivities to certain fluorescence quenchers: the brominated phospholipid 1,2-bis(9,10-dibromostearoyl)-sn-glycero(3)phosphocholine, the calcium ionophore calcimycin (A23187) and its brominated analog (4-bromo-A23187), and the nucleotide analog 2'(3')-O-(2,4,6-trinitrophenyl)-adenosine 5'-triphosphate. We show that tryptophans located at the protein-lipid interface are the main contributors to the well-known fluorescence intensity change occurring in parallel with the conformational rearrangement induced by addition of calcium to the ATPase or its removal; Trp-794 on the ATPase chain may be one of these tryptophans. We also show that tryptophans more deeply embedded in the transmembrane protein structure contribute to the fluorescence change observed upon phosphorylation from inorganic phosphate of the calcium-free ATPase. This phosphorylation step involves opposite changes in the fluorescence quantum yield of tryptophans located in the membrane and in the cytoplasmic regions of the ATPase. This result is in agreement with models in which phosphorylation from inorganic phosphate not only changes the ATPase conformation locally around the catalytic center, but also reorganizes the membrane portion of the ATPase by long-range action, allowing, for instance, the calcium sites to become accessible from the luminal medium.
de Foresta
B
B
Centre de Génétique Moléculaire, Laboratoire propre du Centre National de la Recherche Scientifique associé à l'Université P. et M. Curie, Gif sur Yvette, France.
Champeil
P
P
le Maire
M
M
D000818:Animals; D001692:Biological Transport; D000001:Calcimycin / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D004533:Egtazic Acid; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000737:chemistry*
D000001:Calcimycin; D004533:Egtazic Acid; D014364:Tryptophan; D000252:Calcium-Transporting ATPases