The Relationship between Enzyme Conformational Change, Proton Transfer, and Phosphoryl Transfer in β-Phosphoglucomutase

Abstract

Molecular details for the timing and role of proton transfer in phosphoryl transfer reactions are poorly understood. Here, we have combined QM models, experimental NMR measurements, and X-ray structures to establish that the transition of an archetypal phosphoryl transfer enzyme, βPGM, from a very closed near-attack conformation to a fully closed transition state analogue (TSA) conformation triggers both partial proton transfer from the general acid–base residue to the leaving group oxygen and partial dissociation of the transferring phosphoryl group from the leaving group oxygen. Proton transfer continues but is not completed throughout the reaction path of the phosphoryl transfer with the enzyme in the TSA conformation. Moreover, using interacting quantum atoms (IQA) and relative energy gradient (REG) analysis approaches, we observed that the change in the position of the proton and the corresponding increased electrostatic repulsion between the proton and the phosphorus atom provide a stimulus for phosphoryl transfer in tandem with a reduction in the negative charge density on the leaving group oxygen atom. The agreement between solution-phase 19F NMR measurements and equivalent QM models of βPGMWT and βPGMD10N TSA complexes confirms the protonation state of G6P in the two variants, validating the employed QM models. Furthermore, QM model predictions of an AlF4 distortion in response to the proton position are confirmed using high resolution X-ray crystal structures, not only providing additional validation to the QM models but also further establishing metal fluorides as highly sensitive experimental predictors of active-site charge density distributions.

Atoms in molecules in real space: a fertile field for chemical bonding

“…This idea is nicely demonstrated in the complex energetic mechanisms that have been drawn using the REG method for the peptide hydrolysis in the aspartic active site of the HIV-1 protease 207 and the phosphoryl-transfer process in b-phosphoglucomutase. 208 Molecules of biological relevance range from those formed by a few atoms (e.g., water, ions, small drugs, etc.) to flexible and large macromolecules like proteins and nucleic acids.…”

Hydroboration of imines: intermolecularvs.intramolecular hydride transfer

“…IQA 13 is an energy decomposition procedure which was successfully applied on diverse systems, [14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30] whereas REG approach 31 enables identification of IQA energy components that drive the studied process. [31][32][33][34][35][36][37][38][39][40][41][42][43] We performed a thorough investigation according to two mechanisms: a rearrangement within a single imine-BH 3 species, and hydride transfer which involves two adducts. Additionally, the influence of substituents' size on the corresponding barrier heights is studied.…”

Essential Functional Interplay of the Catalytic Groups in Acid Phosphatase

“…In addition, the proposed catalytic reaction of the Asp18-containing ecAGP can be compared to the reversible isomerization (Glc1P to d -glucose 6-phosphate) catalyzed by β-phosphoglucomutase (βPGM), which has been characterized extensively both experimentally 37 − 41 and computationally. 37 , 41 − 46 However, we note that the Asp nucleophiles of PGMs and HADs cooperate in catalysis with a Mg 2+ cofactor ( Figure 1 c) that is essential in providing electrostatic stabilization. Difference in the electrostatic environment between free and Mg 2+ -paired Asp may limit the mechanistic analogy regarding the function as catalytic nucleophile.…”

“…Chemically ,− and structurally, the ionized aspartate represents an excellent substitute of the histidine for function as the catalytic nucleophile under neutral pH conditions. In addition, the proposed catalytic reaction of the Asp18-containing ecAGP can be compared to the reversible isomerization (Glc1P to d -glucose 6-phosphate) catalyzed by β-phosphoglucomutase (βPGM), which has been characterized extensively both experimentally − and computationally. ,− However, we note that the Asp nucleophiles of PGMs and HADs cooperate in catalysis with a Mg 2+ cofactor (Figure c) that is essential in providing electrostatic stabilization. Difference in the electrostatic environment between free and Mg 2+ -paired Asp may limit the mechanistic analogy regarding the function as catalytic nucleophile.…”