Dataset Information

Structural basis for substrate selectivity and nucleophilic substitution mechanisms in human adenine phosphoribosyltransferase catalyzed reaction.

ABSTRACT: The reversible adenine phosphoribosyltransferase enzyme (APRT) is essential for purine homeostasis in prokaryotes and eukaryotes. In humans, APRT (hAPRT) is the only enzyme known to produce AMP in cells from dietary adenine. APRT can also process adenine analogs, which are involved in plant development or neuronal homeostasis. However, the molecular mechanism underlying substrate specificity of APRT and catalysis in both directions of the reaction remains poorly understood. Here we present the crystal structures of hAPRT complexed to three cellular nucleotide analogs (hypoxanthine, IMP, and GMP) that we compare with the phosphate-bound enzyme. We established that binding to hAPRT is substrate shape-specific in the forward reaction, whereas it is base-specific in the reverse reaction. Furthermore, a quantum mechanics/molecular mechanics (QM/MM) analysis suggests that the forward reaction is mainly a nucleophilic substitution of type 2 (SN2) with a mix of SN1-type molecular mechanism. Based on our structural analysis, a magnesium-assisted SN2-type mechanism would be involved in the reverse reaction. These results provide a framework for understanding the molecular mechanism and substrate discrimination in both directions by APRTs. This knowledge can play an instrumental role in the design of inhibitors, such as antiparasitic agents, or adenine-based substrates.

SUBMITTER: Ozeir M

PROVIDER: S-EPMC6690681 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Structural basis for substrate selectivity and nucleophilic substitution mechanisms in human adenine phosphoribosyltransferase catalyzed reaction.

Ozeir Mohammad M Huyet Jessica J Burgevin Marie-Claude MC Pinson Benoît B Chesney Françoise F Remy Jean-Marc JM Siddiqi Abdul Rauf AR Lupoli Roland R Pinon Grégory G Saint-Marc Christelle C Gibert Jean-François JF Morales Renaud R Ceballos-Picot Irène I Barouki Robert R Daignan-Fornier Bertrand B Olivier-Bandini Anne A Augé Franck F Nioche Pierre P

The Journal of biological chemistry 20190603 32

The reversible adenine phosphoribosyltransferase enzyme (APRT) is essential for purine homeostasis in prokaryotes and eukaryotes. In humans, APRT (hAPRT) is the only enzyme known to produce AMP in cells from dietary adenine. APRT can also process adenine analogs, which are involved in plant development or neuronal homeostasis. However, the molecular mechanism underlying substrate specificity of APRT and catalysis in both directions of the reaction remains poorly understood. Here we present the c ...[more]

PMID: 31160323

Similar Datasets

Project description:In search for the cause leading to low reaction yields, each step along the reaction energy profile computed for the assumed oxidative nucleophilic substitution of hydrogen (ONSH) reaction between 2-phenylquinoxaline and lithium phenylacetylide was modelled computationally. Intermolecular and intramolecular interaction energies and their changes between consecutive steps of ONSH were quantified for molecular fragments playing leading roles in driving the reaction to completion. This revealed that the two reactants have a strong affinity for each other, driven by the strong attractive interactions between Li and two N-atoms, leading to four possible reaction pathways (RP-C2, RP-C3, RP-C5, and RP-C10). Four comparable in energy and stabilizing molecular system adducts were formed, each well prepared for the subsequent formation of a C-C bond at either one of the four identified sites. However, as the reaction proceeded through the TS to form the intermediates (5a-d), very high energy barriers were observed for RP-C5 and RP-C10. The data obtained at the nucleophilic addition stage indicated that RP-C3 was both kinetically and thermodynamically favored over RP-C2. However, the energy barriers observed at this stage were very comparable for both RPs, indicating that they both can progress to form intermediates 5a and 5b. Interestingly, the phenyl substituent (Ph1) on the quinoxaline guided the nucleophile towards both RP-C2 and RP-C3, indicating that the preferred RP cannot be attributed to the steric hindrance caused by Ph1. Upon the introduction of H2O to the system, both RPs were nearly spontaneous towards their respective hydrolysis products (8a and 8b), although only 8b can proceed to the final oxidation stage of the ONSH reaction mechanism. The results suggest that RP-C2 competes with RP-C3, which may lead to a possible mixture of their respective products. Furthermore, an alternative, viable, and irreversible reaction path was discovered for the RP-C2 that might lead to substantial waste. Finally, the modified experimental protocol is suggested to increase the yield of the desired product.

Dataset Information

Structural basis for substrate selectivity and nucleophilic substitution mechanisms in human adenine phosphoribosyltransferase catalyzed reaction.

Publications

Structural basis for substrate selectivity and nucleophilic substitution mechanisms in human adenine phosphoribosyltransferase catalyzed reaction.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets