| Autor: |
Tang L; 1] Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA [2] Howard Hughes Medical Institute, University of Washington, Seattle, Washington 98195, USA [3]., Gamal El-Din TM; 1] Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA [2]., Payandeh J; 1] Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA [2] Department of Structural Biology, Genentech Inc., South San Francisco, California 94080, USA., Martinez GQ; Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA., Heard TM; Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA., Scheuer T; Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA., Zheng N; 1] Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA [2] Howard Hughes Medical Institute, University of Washington, Seattle, Washington 98195, USA., Catterall WA; Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA. |
| Abstrakt: |
Voltage-gated calcium (CaV) channels catalyse rapid, highly selective influx of Ca(2+) into cells despite a 70-fold higher extracellular concentration of Na(+). How CaV channels solve this fundamental biophysical problem remains unclear. Here we report physiological and crystallographic analyses of a calcium selectivity filter constructed in the homotetrameric bacterial NaV channel NaVAb. Our results reveal interactions of hydrated Ca(2+) with two high-affinity Ca(2+)-binding sites followed by a third lower-affinity site that would coordinate Ca(2+) as it moves inward. At the selectivity filter entry, Site 1 is formed by four carboxyl side chains, which have a critical role in determining Ca(2+) selectivity. Four carboxyls plus four backbone carbonyls form Site 2, which is targeted by the blocking cations Cd(2+) and Mn(2+), with single occupancy. The lower-affinity Site 3 is formed by four backbone carbonyls alone, which mediate exit into the central cavity. This pore architecture suggests a conduction pathway involving transitions between two main states with one or two hydrated Ca(2+) ions bound in the selectivity filter and supports a 'knock-off' mechanism of ion permeation through a stepwise-binding process. The multi-ion selectivity filter of our CaVAb model establishes a structural framework for understanding the mechanisms of ion selectivity and conductance by vertebrate CaV channels. |
