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[摘要]：The sarcoplasmic reticulum Ca2+-ATPase transports two Ca2+ per ATP hydrolyzed from the cytoplasm to the lumen against a large concentration gradient. During transport, the pump alters the affinity and accessibility for Ca2+ by rearrangements of transmembrane helices. In this study, all-atom molecular dynamics simulations were performed for wild-type Ca2+-ATPase in the Ca2+-bound form and the Gln mutants of Glu771 and Glu908. Both of them contribute only one carboxyl oxygen to site I Ca2+, but only Glu771Gln completely looses the Ca2+-binding ability. The simulations show that: (i) For Glu771Gln, but not Glu908Gln, coordination of Ca2+ was critically disrupted. (ii) Coordination broke at site II first, although Glu771 and Glu908 only contribute to site I. (iii) A water molecule bound to site I Ca2+ and hydrogen bonded to Glu771 in wild-type, drastically changed the coordination of Ca2+ in the mutant. (iv) Water molecules flooded the binding sites from the lumenal side. (v) The side chain conformation of Ile775, located at the head of a hydrophobic cluster near the lumenal surface, appears critical for keeping out bulk water. Thus the simulations highlight the importance of the water molecule bound to site I Ca2+ and point to a strong relationship between Ca2+-coordination and shielding of bulk water, providing insights into the mechanism of gating of ion pathways in cation pumps. |
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