Quantum Mechanical/Molecular Mechanical Analysis of the Catalytic Mechanism of Phosphoserine Phosphatase.

Dieter Krachtus,Jeremy Smith,Petra Imhof

doi:10.3390/molecules23123342

Abstract

Phosphoserine phosphatase (PSP), a member of the haloacid dehalogenase (HAD) superfamily that comprises the vast majority of phosphotransferases, is likely a steady-state regulator of the level of d-serine in the brain. The proposed catalytic cycle of PSP consists of a two-step mechanism: formation of a phospho-enzyme intermediate by phosphate transfer to Asp11 and its subsequent hydrolysis. Our combined quantum mechanical/molecular mechanical (QM/MM) calculations of the reaction pathways favour a dissociative mechanism of nucleophilic substitution via a trigonal-planar metaphosphate-like configuration for both steps, associated with proton transfer to the leaving group or from the nucleophile. This proton transfer is facilitated by active site residue Asp13 that acts as both a general base and a general acid. Free energy calculation on the reaction pathways further support the structural role of the enzymatic environment and the active site architecture. The choice of a proper reaction coordinate along which to bias the free energy calculations can be guided by a projection of the canonical reaction coordinate obtained from a chain-of-state optimisation onto important internal coordinates.

Highlights

Enzymatic phosphoryl group transfer is found at the core of many cellular biochemical reactions [1]
To evaluate the QM method to be used in the quantum mechanical/molecular mechanical (QM/MM) calculations, purely quantum mechanical calculations of minimal active site models were performed for the two steps of the phosphate transfer reaction in PSP
In the QM/MM study by Re at al. [53], a dissociative pathway is found for Step 1 of the phosphoryl transfer mechanism and in our present computations, the mechanisms of both Step 1 and Step 2, are clearly dissociative: As observed in the previous study on Step 1 [53], the meta-phosphate and oxygen atoms of the nucleophile and the leaving group are in apical positions resembling a trigonal-bipyramidal associative transition state, similar to that mimicked in the crystal structures of the PSP-BeF3 and PSP-AlF3 complexes [26]

Summary

Introduction

Enzymatic phosphoryl group transfer is found at the core of many cellular biochemical reactions [1]. The consistently low rate of this reaction in aqueous solution [6] is enhanced by up to ∼1021 by the catalytic action of phosphotransferases These enzymes lower the kinetic barrier through general acid–base catalysis, nucleophilic catalysis and by strongly stabilizing (Kd ∼10−26 M) the reaction intermediates [6] through an ensemble of electrostatic and steric interactions [7,8,9]. The specificity of these interactions is vital for phosphoryl group transfer as exemplified by the independent co-evolution of a strikingly similar catalytic scaffold. Molecules 2018, 23, 3342; doi:10.3390/molecules23123342 www.mdpi.com/journal/molecules (see Figure 1) in many families of phosphotransferases belonging to the haloacid dehalogenase-like hydrolase (HAD) superfamily [10], comprising phosphatases, phosphomutases, and P-type ATPases.

Methods

Results

Discussion

Conclusion