scholarly journals With or without light: comparing the reaction mechanism of dark-operative protochlorophyllide oxidoreductase with the energetic requirements of the light-dependent protochlorophyllide oxidoreductase

2014 ◽  
Author(s):  
Pedro J Silva
Keyword(s):  
Reaction Mechanism ◽  
Electron Donor ◽  
Ph Dependence ◽  
Single Point ◽  
Enzyme Reaction ◽  
Single Electron ◽  
Simple Explanation ◽  
Dependent Manner ◽  
Protochlorophyllide Oxidoreductase ◽  
Sequence Of Events

The addition of two electrons and two protons to the C17=C18 bond in protochlorophyllide is catalyzed by a light-dependent enzyme relying on NADPH as electron donor, and by a light-independent enzyme bearing a (Cys)3Asp-ligated [4Fe-4S] cluster which is reduced by cytoplasmic electron donors in an ATP-dependent manner and then functions as electron donor to protochlorophyllide. The precise sequence of events occurring at the C17=C18 bond has not, however, been determined experimentally in the dark-operating enzyme. In this paper, we present the computational investigation of the reaction mechanism of this enzyme at the B3LYP/6-311+G(d,p)// B3LYP/6-31G(d) level of theory. The reaction mechanism begins with single-electron reduction of the substrate by the (Cys)3Asp-ligated [4Fe-4S], yielding a negatively-charged intermediate. Depending on the rate of Fe-S cluster re-reduction, the reaction either proceeds through double protonation of the single-electron-reduced substrate, or by alternating proton/electron transfer. The computed reaction barriers suggest that Fe-S cluster re-reduction should be the rate-limiting stage of the process. Poisson-Boltzmann computations on the full enzyme-substrate complex, followed by Monte Carlo simulations of redox and protonation titrations revealed a hitherto unsuspected pH-dependence of the reaction potential of the Fe-S cluster. Furthermore, the computed distributions of protonation states of the His, Asp and Glu residues were used in conjunction with single-point ONIOM computations to obtain, for the first time, the influence of all protonation states of an enzyme on the reaction it catalyzes. Despite exaggerating the ease of reduction of the substrate, these computations confirmed the broad features of the reaction mechanism obtained with the medium-sized models, and afforded valuable insights on the influence of the titratable amino acids on each reaction step. Additional comparisons of the energetic features of the reaction intermediates with those of common biochemical redox intermediates suggest a surprisingly simple explanation for the mechanistic differences between the dark-catalyzed and light-dependent enzyme reaction mechanisms.

scholarly journals With or without light: comparing the reaction mechanism of dark-operative protochlorophyllide oxidoreductase with the energetic requirements of the light-dependent protochlorophyllide oxidoreductase

2014 ◽  
Author(s):  
Pedro J Silva
Keyword(s):  
Reaction Mechanism ◽  
Electron Donor ◽  
Ph Dependence ◽  
Single Point ◽  
Enzyme Reaction ◽  
Single Electron ◽  
Simple Explanation ◽  
Dependent Manner ◽  
Protochlorophyllide Oxidoreductase ◽  
Sequence Of Events

The addition of two electrons and two protons to the C17=C18 bond in protochlorophyllide is catalyzed by a light-dependent enzyme relying on NADPH as electron donor, and by a light-independent enzyme bearing a (Cys)3Asp-ligated [4Fe-4S] cluster which is reduced by cytoplasmic electron donors in an ATP-dependent manner and then functions as electron donor to protochlorophyllide. The precise sequence of events occurring at the C17=C18 bond has not, however, been determined experimentally in the dark-operating enzyme. In this paper, we present the computational investigation of the reaction mechanism of this enzyme at the B3LYP/6-311+G(d,p)// B3LYP/6-31G(d) level of theory. The reaction mechanism begins with single-electron reduction of the substrate by the (Cys)3Asp-ligated [4Fe-4S], yielding a negatively-charged intermediate. Depending on the rate of Fe-S cluster re-reduction, the reaction either proceeds through double protonation of the single-electron-reduced substrate, or by alternating proton/electron transfer. The computed reaction barriers suggest that Fe-S cluster re-reduction should be the rate-limiting stage of the process. Poisson-Boltzmann computations on the full enzyme-substrate complex, followed by Monte Carlo simulations of redox and protonation titrations revealed a hitherto unsuspected pH-dependence of the reaction potential of the Fe-S cluster. Furthermore, the computed distributions of protonation states of the His, Asp and Glu residues were used in conjunction with single-point ONIOM computations to obtain, for the first time, the influence of all protonation states of an enzyme on the reaction it catalyzes. Despite exaggerating the ease of reduction of the substrate, these computations confirmed the broad features of the reaction mechanism obtained with the medium-sized models, and afforded valuable insights on the influence of the titratable amino acids on each reaction step. Additional comparisons of the energetic features of the reaction intermediates with those of common biochemical redox intermediates suggest a surprisingly simple explanation for the mechanistic differences between the dark-catalyzed and light-dependent enzyme reaction mechanisms.

scholarly journals With or without light: comparing the reaction mechanism of dark-operative protochlorophyllide oxidoreductase with the energetic requirements of the light-dependent protochlorophyllide oxidoreductase

2014 ◽  
Author(s):  
Pedro J Silva
Keyword(s):  
Reaction Mechanism ◽  
Electron Donor ◽  
Enzyme Reaction ◽  
Single Electron ◽  
Simple Explanation ◽  
Dependent Manner ◽  
Protochlorophyllide Oxidoreductase ◽  
Sequence Of Events ◽  
Limiting Stage ◽  
Reaction Barriers

The addition of two electrons and two protons to the C17=C18 bond in protochlorophyllide is catalyzed by a light-dependent enzyme relying on NADPH as electron donor, and by a light-independent enzyme bearing a (Cys)3Asp-ligated [4Fe-4S] cluster which is reduced by cytoplasmic electron donors in an ATP-dependent manner and then functions as electron donor to protochlorophyllide. The precise sequence of events occurring at the C17=C18 bond has not, however, been determined experimentally in the dark-operating enzyme. In this paper, we present the computational investigation of the reaction mechanism of this enzyme at the B3LYP/6-311+G(d,p)// B3LYP/6-31G(d) level of theory. The reaction mechanism begins with single-electron reduction of the substrate by the (Cys)3Asp-ligated [4Fe-4S], yielding a negatively-charged intermediate. Depending on the rate of Fe-S cluster re-reduction, the reaction either proceeds through double protonation of the single-electron-reduced substrate, or by alternating proton/electron transfer The computed reaction barriers suggest that Fe-S cluster re-reduction should be the rate-limiting stage of the process. Additional comparisons of the energetic features of the intermediates with those of common biochemical redox intermediates suggest a surprisingly simple explanation for the mechanistic differences between the dark-catalyzed and light-dependent enzyme reaction mechanisms.

Rationalizing the pH-Activity Response for Escherichia Coli’s Glycinamide Ribonucleotidetransformylase Through Computational Methods

2019 ◽  
Author(s):  
Adrian Roitberg ◽  
Pancham Lal Gupta
Keyword(s):  
Transfer Reaction ◽  
Catalytic Mechanism ◽  
Ph Dependence ◽  
Ph Effects ◽  
Dependent Manner ◽  
E Coli ◽  
Gar Tfase ◽  
Activity Curve ◽  
Ph Dependent ◽  
Formyl Transfer

<div>Human Glycinamide ribonucleotide transformylase (GAR Tfase), a regulatory enzyme in the de novo purine biosynthesis pathway, has been established as an anti-cancer target. GAR Tfase catalyzes the formyl transfer reaction from the folate cofactor to the GAR ligand. In the present work, we study E. coli GAR Tfase, which has high sequence similarity with the human GAR Tfase with most functional residues conserved. E. coli GAR Tfase exhibits structural changes and the binding of ligands that varies with pH which leads to change the rate of the formyl transfer reaction in a pH-dependent manner. Thus, the inclusion of pH becomes essential for the study of its catalytic mechanism. Experimentally, the pH-dependence of the kinetic parameter kcat is measured to evaluate the pH-range of enzymatic activity. However, insufficient information about residues governing the pH-effects on the catalytic activity leads to ambiguous assignments of the general acid and base catalysts and consequently its catalytic mechanism. In the present work, we use pH-replica exchange molecular dynamics (pH-REMD) simulations to study the effects of pH on E. coli GAR Tfase enzyme. We identify the titratable residues governing the pH-dependent conformational changes in the system. Furthermore, we filter out the protonation states which are essential in maintaining the structural integrity, keeping the ligands bound and assisting the catalysis. We reproduce the experimental pH-activity curve by computing the population of key protonation states. Moreover, we provide a detailed description of residues governing the acidic and basic limbs of the pH-activity curve.</div>

scholarly journals Calcium-sensitive, lipid-binding cytoskeletal proteins of the human placental microvillar region.

1987 ◽  
Vol 105 (1) ◽  
pp. 303-311 ◽  
Author(s):  
H C Edwards ◽  
A G Booth
Keyword(s):  
Lipid Binding ◽  
Cytoskeletal Proteins ◽  
Dependent Manner ◽  
Molecular Weights ◽  
Sequence Of Events ◽  
Marked Preference ◽  
High Concentrations

In this study we describe a group of Ca2+-sensitive proteins located in the microvillar region of the human placental syncytiotrophoblast. By following the distribution of proteins between the particulate and supernatant phases of detergent-solubilized microvilli in the presence of defined concentrations of free Ca2+, we demonstrate a class of proteins of subunit molecular weights 72,000, 69,000, 38,000, 36,000, and 32,000 that associate with both the cytoskeleton and lipid at high concentrations of free Ca2+. These proteins can be released from microvilli using EGTA-containing buffers. Although they do not bind to phenyl-Sepharose, they will bind to phospholipids immobilized on phenyl-Sepharose columns in a Ca2+-dependent manner and show a marked preference for phospholipids with negatively charged headgroups. The results provide evidence for a sequence of events which may occur within the microvillus as the localized concentration of intracellular free Ca2+ rises.

scholarly journals KOtBu as a Single Electron Donor? Revisiting the Halogenation of Alkanes with CBr4 and CCl4

Molecules ◽  
2018 ◽  
Vol 23 (5) ◽  
pp. 1055 ◽  
Author(s):  
Katie Emery ◽  
Allan Young ◽  
J. Arokianathar ◽  
Tell Tuttle ◽  
John Murphy
Keyword(s):  
Electron Donor ◽  
Single Electron

scholarly journals The effect of disulfiram on the aldehyde dehydrogenases of sheep liver

1975 ◽  
Vol 151 (2) ◽  
pp. 407-412 ◽  
Author(s):  
T M Kitson
Keyword(s):  
Ph Dependence ◽  
Maximum Velocity ◽  
Enzyme Reaction ◽  
Thiol Group ◽  
Inhibitory Effect ◽  
Mitochondrial Enzyme ◽  
Aldehyde Dehydrogenases ◽  
Sheep Liver ◽  
Rate Profile

1. The effect of disulfiram on the activity of the cytoplasmic and mitochondrial aldehyde dehydrogenases of sheep liver was studied. 2. Disulfiram causes an immediate inhibition of the enzyme reaction. The effect on the cytoplasmic enzyme is much greater than on the mitochondrial enzyme. 3. In both cases, the initial partial inhibition is followed by a gradual irreversible loss of activity. 4. The pH-rate profile of the inactivation of the mitochondrial enzyme by disulfiram and the pH-dependence of the maximum velocity of the enzyme-catalysed reaction are both consistent with the involvement of a thiol group. 5. Excess of 2-mercaptoethanol or GSH abolishes the effect of disulfiram. However, equimolar amounts of either of these reagents and disulfiram cause an effect greater than does disulfiram alone. It was shown that the mixed disulphide, Et2N-CS-SS-CH2-CH2OH, strongly inhibits aldehyde dehydrogenase. 6. The inhibitory effect of diethyldithiocarbamate in vitro is due mainly to contamination by disulfiram.

Theoretical study on the OH-initiated atmospheric reaction of N-methyl perfluorobutane sulfonamidoethanol (C4F9SO2N(CH3)CH2CH2OH)

2013 ◽  
Vol 91 (12) ◽  
pp. 1161-1167
Author(s):  
Juan Dang ◽  
Lei Ding ◽  
Xiaoyan Sun ◽  
Qingzhu Zhang ◽  
Wenxing Wang
Keyword(s):  
Reaction Mechanism ◽  
Degradation Products ◽  
Single Point ◽  
Basis Set ◽  
Stationary Points ◽  
Geometrical Parameters ◽  
Chemical Transformations ◽  
Point Energy ◽  
Range Transport ◽  
Perfluorinated Carboxylic Acids

N-methyl perfluorobutane sulfonamidoethanol (NMeFBSE), a new product of the 3M Company, is currently widely used in many countries and territories. It is prone to volatilize to the atmosphere where it can undergo long-range transport and chemical transformations. In this work, the reaction mechanism for the OH-initiated atmospheric oxidation of NMeFBSE was investigated. The geometrical parameters and vibrational frequencies of all of the stationary points were calculated at the MPWB1K level with the 6-31G+(d,p) basis set. Single-point energy calculations were carried out at the MPWB1K/6-311+G(3df,2p) level. The results indicate that the channel of the formation of C4F9 and HSO3N(CH3)CH2CH2OH resulting from OH addition to NMeFBSE and hydrogen abstractions from the −CH3 group in NMeFBSE are energetically favorable. The main degradation products include perfluorinated carboxylic acids (C3F7COOH, C2F5COOH, CF3COOH), HSO3N(CH3)CH2CH2OH, NMeFBSA (C4F9SO2NH(CH3)), C4F9SO2N(CH3)CH2CHO, and C4F9SO2N(CH3)CH2COOH. The reaction mechanism for the formation of NMeFBSA is reported for the first time. Using the atmospheric fate of NMeFBSE as a guide, it seems that N-methyl perfluorooctane sulfonamidoethanol (NMeFOSE) contributes to the ubiquity of perfluoroalkyl sulfonate and carboxylate compounds in the atmosphere.

Comparing lactate and glycerol as a single-electron donor for sulfate reduction in fluidized bed reactors

2014 ◽  
Vol 25 (5) ◽  
pp. 719-733 ◽  
Author(s):  
Sueli M. Bertolino ◽  
Lucas A. Melgaço ◽  
Renata G. Sá ◽  
Versiane A. Leão
Keyword(s):  
Sulfate Reduction ◽  
Fluidized Bed ◽  
Electron Donor ◽  
Single Electron ◽  
Fluidized Bed Reactors
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