DFT studies on key intermediates in thiamin catalysis

Abstract

ABSTRACT:The diphosphate ester (ThDP) of thiamin (vitamin B 1 ) is an important cofactor of enzymes within the carbohydrate metabolism. From experiments of site-specific variants and nuclear magnetic resonance (NMR) studies, it is known that the protonation of the N1Ј atom is a significant step in the coenzyme activation by the enzymatic environment. Therefore, we have performed density functional theory (DFT) calculations on the B3LYP/ 6-31G* level of N1ЈH and N1ЈCH 3 thiamin as model systems to study the protonation and methylation effect on the structure and the electronic properties of the 4Ј-amino group. The relaxed rotational barriers related to the C4Ј-4ЈN bond are correlated with findings of 1 H NMR studies and proton/deuterium exchange experiments. Moreover, the effect of N1Ј protonation was studied in more detail on the hydroxyethyl-thiamin carbanion (HETh Ϫ ), a key intermediate during catalysis of some ThDP-dependent enzymes. The relaxed rotational barriers related to the C2OC2␣ bond and the reaction coordinates of the proton transfer 4ЈNOH3 C2␣ of HETh Ϫ and N1ЈH-HETh Ϫ show that they are significantly determined by the protonation at N1Ј of HETh Ϫ . The influence of the apoenzyme environment on the active coenzyme conformation is modeled in a very simple way. The characteristic torsion angles ⌽ T and ⌽ P are considered to be restricted in terms of their values in the corresponding enzyme as well as free optimization parameters. Frequency calculations were performed to characterize the minima and transition state structures, respectively. The applicability of the DFT method was checked by comparing calculations on the MP2-HF-SCF/6-31G* level.

Computational characterization of enzyme-bound thiamin diphosphate reveals a surprisingly stable tricyclic state: implications for catalysis

“…For example, in some very early work, Jordan used semi-empirical methods to study the electronic structure and conformational space of the cofactor in the gas phase, acknowledging the difficulty of comparing these results to reactions in solution and on the enzyme [19–20]. Thirty years later, density functional theory (DFT) calculations showed that the 4′-amino moiety of the cofactor can either accept or donate a proton in the reactions, depending on the protonation state of N1′ [21].…”

A Theoretical Study of the Benzoylformate Decarboxylase Reaction Mechanism

“…The adopted methodology, called the cluster approach, has been very successful in elucidating reaction mechanisms of a wide variety of enzymes (Siegbahn and Himo, 2009 , 2011 ; Siegbahn and Blomberg, 2010 ; Blomberg et al, 2014 ; Himo, 2017 ). A number of previous theoretical studies using a variety of computational techniques have considered aspects of mechanism of a number of ThDP-dependent enzymes (Friedemann et al, 2004 ; Lie et al, 2005 ; Wang et al, 2005 ; Jaña et al, 2010 ; Topal et al, 2010 ; Xiong et al, 2010 ; Paramasivam et al, 2011 ; Alvarado et al, 2012 ; Hou et al, 2012 ; Jaña and Delgado, 2013 ; Sheng and Liu, 2013 ; Sheng et al, 2013 , 2014 ; Sánchez et al, 2014 ; Zhang et al, 2014 ; Zhu and Liu, 2014 ; Lizana et al, 2015 ; Nauton et al, 2016 ; White et al, 2016 ). Relevant results from these studies have been incorporated into the discussion of the BFDC mechanism below.…”

Computational screening of novel thiamine-catalyzed decarboxylation reactions of 2-keto acids

“…Experimental [30] and theoretical studies [31][32][33][34][35] have been aimed at understanding the mechanism of the reaction and the structures of the intermediates associated with the reaction. The first step of the pathway can be written as Pyruvic acid þ CoA þ ferredoxin ðoxÞ $ acetyl-CoA þ CO 2 þ ferredoxin ðredÞ ð 1Þ…”

“…The intermediate (C) undergoes an internal proton transfer via a barrier (D, TS2) to form 2-lactyl ThDP (E, LThDP).Decarboxylation of LThDP occurs through a transition state (F, TS3) to form a stable enamine and CO 2 (G). Detailed experimental studies have been published that delineate the complete decarboxylation process of pyruvate dehydrogenases (PDH), pyruvate decarboxylases (PDC) and pyruvate oxidases (POX) by various research groups[2,30,34,60,[66][67][68][69][70].…”