Comparative analysis of active sites in P-loop nucleoside triphosphatases suggests an ancestral activation mechanism

Abstract

P-loop nucleoside triphosphatases (NTPases) share common Walker A (P-loop) and Walker B sequence motifs and depend on activating moieties (Arg or Lys fingers or a K+ ion). In search for a common catalytic mechanism, we combined structure comparisons of active sites in major classes of P-loop NTPases with molecular dynamics (MD) simulations of the Ras GTPase, a well-studied oncoprotein. Comparative structure analysis showed that positively charged activating moieties interact with gamma-phosphate groups of NTP substrates in all major classes of P-loop NTPases. In MD simulations, interaction of the activating Arg finger with the Mg-GTP-Ras complex led to the rotation of the gamma-phosphate group by 40 degrees enabling its interaction with the backbone amide group of Gly13. In all analyzed structures, the residue that corresponds to Gly13 of Ras was in a position to stabilize gamma-phosphate after its rotation, suggesting a common ancestral activation mechanism within the entire superfamily.