Interdomain communication in calcium pump as revealed in the crystal structures with transmembrane inhibitors.
Ca(2+)-ATPase of skeletal muscle sarcoplasmic reticulum is an ATP-driven Ca(2+) pump consisting of three cytoplasmic domains and 10 transmembrane helices. In the absence of Ca(2+), the three cytoplasmic domains gather to form a compact headpiece, but the ATPase is unstable without an inhibitor. Here we describe the crystal structures of Ca(2+)-ATPase in the absence of Ca(2+) stabilized with cyclopiazonic acid alone and in combination with other inhibitors. Cyclopiazonic acid is located in the transmembrane region of the protein near the cytoplasmic surface. The binding site partially overlaps with that of 2,5-di-tert-butyl-1,4-dihydroxybenzene but is separate from that of thapsigargin. The overall structure is significantly different from that stabilized with thapsigargin: The cytoplasmic headpiece is more upright, and the transmembrane helices M1-M4 are rearranged. Cyclopiazonic acid primarily alters the position of the M1' helix and thereby M2 and M4 and then M5. Because M5 is integrated into the phosphorylation domain, the whole cytoplasmic headpiece moves. These structural changes show how an event in the transmembrane domain can be transmitted to the cytoplasmic domain despite flexible links between them. They also reveal that Ca(2+)-ATPase has considerable plasticity even when fixed by a transmembrane inhibitor, presumably to accommodate thermal fluctuations.
Takahashi
Mihoko
M
Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan.
Kondou
Youhei
Y
Toyoshima
Chikashi
C
D000818:Animals; D001665:Binding Sites; D018360:Crystallography, X-Ray; D003593:Cytoplasm / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006860:Hydrogen Bonding; D007211:Indoles / Q000494:pharmacology*; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008956:Models, Chemical; D008958:Models, Molecular; D011485:Protein Binding; D017433:Protein Structure, Secondary / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000187:drug effects; D012519:Sarcoplasmic Reticulum / Q000201:enzymology; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D019284:Thapsigargin / Q000378:metabolism / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007211:Indoles; D008565:Membrane Proteins; D019284:Thapsigargin; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; C000543:cyclopiazonic acid