Computer modelling reveals new conformers of the ATP binding loop of Na+/K+-ATPase involved in the transphosphorylation process of the sodium pump
Abstract
Hydrolysis of ATP by Na+/K+-ATPase, a P-Type ATPase, catalyzing active Na+ and K+ transport through cellular membranes leads transiently to a phosphorylation of its catalytical alpha-subunit. Surprisingly, three-dimensional molecular structure analysis of P-type ATPases reveals that binding of ATP to the N-domain connected by a hinge to the P-domain is much too far away from the Asp(369) to allow the transfer of ATP's terminal phosphate to its aspartyl-phosphorylation site.
In order to get information for how the transfer of the gamma-phosphate group of ATP to the Asp(369) is achieved, analogous molecular modeling of the M-4 M-5 loop of ATPase was performed using the crystal data of Na+/K+-ATPase of different species. Analogous molecular modeling of the cytoplasmic loop between Thr(338) and Ile(760) of the a alpha-subunit of Na+/K+-ATPase and the analysis of distances between the ATP binding site and phosphorylation site revealed the existence of two ATP binding sites in the open conformation; the first one close to Phe(475) in the N-domain, the other one close to Asp(369) in the 13-domain.
However, binding of Mg2+circle ATP to any of these sites in the "open conformation" may not lead to phosphorylation of Asp(369). Additional conformations of the cytoplasmic loop were found wobbling between "open conformation" "semi-open conformation "closed conformation" in the absence of 2Mg(2+)circle ATP.
The cytoplasmic loop's conformational change to the "semi-open conformation" characterized by a hydrogen bond between Arg(543) and Asp(611) triggers by binding of 2Mg(2+)circle ATP Ito a single ATP l site and conversion to the "closed l conformation" the phosphorylation of Asp(369) in the P-domain, and hence the start of Na+/K+-activated ATP hydrolysis.