The beryllium fluoride form of the Na+,K+-ATPase and the intermediates of the functional cycle

The Na+,K+-ATPase generates electrochemical gradients of Na+ and K+ across the plasma membrane. Here, we describe a 4.0 Å resolution crystal structure of the pig kidney Na+,K+-ATPase stabilized by beryllium fluoride (denoted E2-BeFx). The structure shows high resemblance to the E2P phosphoenzyme obtained by phosphorylation from inorganic phosphate (Pi) and stabilised by cardiotonic steroids, and reveals a Mg2+ bound near the ion binding site II. Anomalous Fourier analysis of the crystals soaked in Rb+ (K+ congener) followed by a low resolution rigid-body refinement (6.9-7.5 Å) revealed pre-occlusion transitions leading to activation of the desphosphorylation reaction. Mg2+ location indicates a site of an initial K+ recognition and acceptance upon binding to the outward-open E2P state after Na+ release. Despite the overall structural resemblance to the Pi-induced E2P phosphoform, BeFx inhibited enzyme is able to binds both ADP/ATP and ions, the features that relate E2-BeFx complex to an intermediate of the functional cycle of the Na+,K+-ATPase prior E2P.

Identification of beryllium fluoride complexes in mechanically distorted gels using quadrupolar split 9Be NMR spectra resolved withsolution-state selective cross-polarization

The uniformly anisotropic media afforded by hydrogels are being increasingly exploited in analytical (structural) nuclear magnetic The uniformly anisotropic media afforded by hydrogels are being increasingly exploited in analytical (structure...