Common mechanism of activated catalysis in P-loop fold nucleoside triphosphatases -in varietate concordia

Abstract

Although P-loop fold nucleoside triphosphatases (also known as Walker NTPases) are ubiquitous, their catalytic mechanism remains obscure. Based on a comparative structural analysis of 3136 Mg-NTP-containing catalytic sites, we propose a common scheme of activated catalysis for P-loop NTPases where a hydrogen bond (H-bond) between the strictly conserved, Mg-coordinating Ser/Thr of the Walker A motif ([Ser/Thr]WA) and the conserved aspartate of the Walker B motif (AspWB) plays the key role. We found that this H-bond is very short in the structures with bound transition state (TS) analogs. We suggest that the proton affinities of these two residues reverse in the TS so that the proton relocates from [Ser/Thr]WA to AspWB. The anionic [Ser/Thr]WA withdraws then a proton from the (catalytic) water molecule, and the nascent hydroxyl anion attacks gamma-phosphate. When the gamma-phosphate group breaks away, the trapped proton relays from AspWB, via [Ser/Thr]WA, to beta-phosphate and compensates for its developing negative charge.

Common patterns of hydrolysis initiation in P-loop fold nucleoside triphosphatases

“…In such complexes, a water molecule is often located apically to the analog of γ-phosphate, which corroborates the earlier suggestions that γ-phosphate cleavage is triggered by the apical nucleophilic attack of a polarized water molecule (Wcat), after its deprotonation to OHcat, on γ-phosphate, see Fig. 1C, D; the accompanying article [18] and [12,35,[44][45][46][47][48][49]. Also, TS analogs were shown to promote the interaction of the activator with the NTPase domain and therefore often disclose the interaction of the stimulator with the triphosphate chain, as shown in Fig.…”

“…The bending of the triphosphate chain of a P-loop-bound NTP molecule is not possible because of its multiple bonds with the protein, see Fig. 1C, D, the accompanying article [18] and [70]. At the same time, the examination of available structures revealed several cases which can be considered as evidence of γ-phosphate twisting in the TS (see Fig.…”

“…We provide the relevant outlook on the future studies of P-loop NTPases in the last section of the accompanying paper [18]. Here, instead, we would like to give a flashback.…”

“…The main classes of P-loop NTPases, which are listed in Fig. S1 of Supplementary Figures and characterized in the accompanying article [18], were already present in the Last Universal Cellular Ancestor (LUCA) [7,8,13,15,[19][20][21][22][23].…”

“…This common feature allowed us to suggest that stimulators may trigger NTP hydrolysis by displacing the oxygen atoms of γ-phosphate, in particular, the O 1G atom that coordinates the Mg 2+ ion. In the accompanying article [18] we further consider the functional consequences of the γ-phosphate movement and suggest a common catalytic mechanism for P-loop fold NTPases.…”