共查询到20条相似文献,搜索用时 31 毫秒
1.
Rong-Zhen Liao 《Journal of biological inorganic chemistry》2013,18(2):175-181
Formaldehyde ferredoxin oxidoreductase is a tungsten-dependent enzyme that catalyzes the oxidative degradation of formaldehyde to formic acid. The molybdenum ion can be incorporated into the active site to displace the tungsten ion, but is without activity. Density functional calculations have been employed to understand the incapacitation of the enzyme caused by molybdenum substitution. The calculations show that the enzyme with molybdenum (Mo-FOR) has higher redox potential than that with tungsten, which makes the formation of the MoVI=O complex endothermic by 14 kcal/mol. Following our previously suggested mechanism for this enzyme, the formaldehyde substrate oxidation was also investigated for Mo-FOR using the same quantum-mechanics-only model, except for the displacement of tungsten by molybdenum. The calculations demonstrate that formaldehyde oxidation occurs via a sequential two-step mechanism. Similarly to the tungsten-catalyzed reaction, the MoVI=O species performs the nucleophilic attack on the formaldehyde carbon, followed by proton transfer in concert with two-electron reduction of the metal center. The first step is rate-limiting, with a total barrier of 28.2 kcal/mol. The higher barrier is mainly due to the large energy penalty for the formation of the MoVI=O species. 相似文献
2.
R D Stebbins E Meller H Rosengarten A Friedhoff R Silber 《Archives of biochemistry and biophysics》1976,173(2):673-679
An enzyme in human platelets or rat brain incubated with 5-methyl tetrahydrofolate (5MeH4folate) yields formaldehyde (4, 13), which will combine with biogenic amines to form β-carbolines (5) or tetrahydroisoquinolines. This activity was purified 500-fold from human platelets which are the main storage site for 5-hydroxytryptamine in man. This enzyme was identical to N5, N10-methylene tetrahydrofolate (N5,N10-methylene H4folate) reductase by the following criteria: (i) co-purification, (ii) heat denaturation, (iii) pH response, (iv) molecular weight, (5) cofactor requirements. A mechanism involving the enzymatic generation of formaldehyde followed by adduct formation with a biogenic amine is proposed. 相似文献
3.
Yan-Fang Liu Rong-Zhen Liao Wan-Jian Ding Jian-Guo Yu Ruo-Zhuang Liu 《Journal of biological inorganic chemistry》2011,16(5):745-752
The reaction mechanism of the hydration of acetylene to acetaldehyde catalyzed by [WIVO(mnt)2]2− (where mnt2− is 1,2-dicyanoethylenedithiolate) is studied using density functional theory. Both the uncatalyzed and the catalyzed reaction
are considered to find out the origin of the catalysis. Three different models are investigated, in which an aquo, a hydroxo,
or an oxo coordinates to the tungsten center. A first-shell mechanism is suggested, similarly to recent calculations on tungsten-dependent
acetylene hydratase. The acetylene substrate first coordinates to the tungsten center in an η2 fashion. Then, the tungsten-bound hydroxide activates a water molecule to perform a nucleophilic attack on the acetylene,
resulting in the formation of a vinyl anion and a tungsten-bound water molecule. This is followed by proton transfer from
the tungsten-bound water molecule to the newly formed vinyl anion intermediate. Tungsten is directly involved in the reaction
by binding and activating acetylene and providing electrostatic stabilization to the transition states and intermediates.
Three other mechanisms are also considered, but the associated energetic barriers were found to be very high, ruling out those
possibilities. 相似文献
4.
Two-layer ONIOM calculations have been carried out to study methanol to propene (MTP) conversion reactions catalyzed by H-beta zeolite. On the basis of the so-called side-chain hydrocarbon pool (HCP) mechanism, this work proposes the complete catalytic cycle pathway for the MTP reaction. The cycle starts from the methylation of pentamethylbenzene (PMB), which leads to the formation of hexamethylbenzenium ion (hexaMB+). Subsequent steps involving deprotonation, methylation, an internal H-shift, and a unimolecular CH3-shift are required to produce propene and ethene. The calculated activation barriers and reaction energy data indicate that propene is the more favored product, rather than ethene, from both kinetic and thermodynamic perspectives, which is consistent with experimental observations. In addition, the calculations suggest that the activation barriers of the reaction steps decrease in the order: internal H-shift?>?methylation?>?unimolecular CH3-shift?≥?deprotonation. In the methylation step, methylation of the exocyclic double bond is easier than methylation of the ring carbons on the aromatic benzene derivative. 相似文献
5.
Ryogo Sugitani 《BBA》2009,1787(9):1140-6960
We have examined the network of connected internal cavities in cytochrome c oxidase along which water produced at the catalytic center is removed from the enzyme. Using combination of structural analysis, molecular dynamics simulations, and free energy calculations we have identified two exit pathways that connect the Mg2+ ion cavity to the outside of the enzyme. Each pathway has a well-defined bottleneck, which determines the overall rate of water traffic along the exit pathway, and a specific cooperative mechanism of passing it. One of the pathways is going via Arg438/439 (in bovine numbering) toward the CuA center, approaching closely its His204B ligand and Lys171B residue; and the other is going toward Asp364 and Thr294. Comparison of the pathways among different aa3-type enzymes shows that they are well conserved. Possible connections of the finding to redox-coupled proton pumping mechanism are discussed. We propose specific mutations near the bottlenecks of the exit pathways that can test some of our hypotheses. 相似文献
6.
Gerard Berger Guy Girault Jean-Michel Galmiche Stephane Pezennec 《Journal of bioenergetics and biomembranes》1994,26(3):335-346
The influences of total magnesium ion concentration at different total ATP concentrations, and of total ATP concentration, for different total magnesium ion concentrations, on the enzymatic rate of the isolated chloroplast F1 ATPase, have been followed by a chromatographic method consisting in the separation and determination of ADP. From the various series of curves, it is concluded that the experimental results (position of the maxima,K
m
values) are better fitted by a mechanism involving the activation of the enzyme by magnesium ion and hydrolysis of free ATP, rather than by the classical mechanism, for which the enzyme hydrolyzes the MgATP complex and is inhibited by Mg2+. Although the equations giving the reaction rate are similar in the two cases, the calculated values ofK
m
are widely different. The value obtained from the classical mechanism does not agree withK
D
, the dissociation constant of the enzyme-substrate complex, measured by the Hummel and Dreyer method. Moreover, when the total ATP concentration tends toward the total magnesium ion concentration, the nucleotide binding to the enzyme tends toward zero, although it should be maximum if MgATP were the true substrate. Finally, the inhibitory effect of Na+ is more easily explained as a competition between this ion and the activating Mg2+, than by the classical mechanism. 相似文献
7.
The mechanism of enzymatic elongation by Escherichia coli DNA polymerase II of a DNA primer, which is annealed to a unique position on the bacteriophage fd viral DNA, has been studied. The enzyme is found to dissociate from the substrate at specific positions on the genome which act as “barriers” to further primer extension. It is believed these are sites of secondary structure in the DNA. When the template is complexed with E. coli DNA binding protein many of these barriers are eliminated and the enzyme remains associated with the same primer-template molecule during extensive intervals of DNA synthesis. Despite the presence of E. coli DNA binding protein, at least one barrier on the fd genome remains rate-limiting to chain extension and disturbs the otherwise processive mechanism of DNA synthesis. This barrier is overcome by increasing the concentration of enzyme.In contrast, it is found that DNA polymerase I is not rate-limited by structural barriers in the template, however, it exhibits a non-processive mechanism of elongation.These findings provide a framework for understanding the necessity for participation of proteins other than a DNA polymerase in chain extension during chromosomal replication. 相似文献
8.
Extracts of Pseudomonas C grown on methanol as sole carbon and energy source contain a methanol dehydrogenase activity which can be coupled to phenazine methosulfate. This enzyme catalyzes two reactions namely the conversion of methanol to formaldehyde (phenazine methosulfate coupled) and the oxidation of formaldehyde to formate (2,6-dichloroindophenol-coupled). Activities of glutathione-dependent formaldehyde dehydrogenase (NAD+) and formate dehydrogenase (NAD+) were also detected in the extracts.The addition of d-ribulose 5-phosphate to the reaction mixtures caused a marked increase in the formaldehyde-dependent reduction of NAD+ or NADP+. In addition, the oxidation of [14C]formaldehyde to CO2, by extracts of Pseudomonas C, increased when d-ribulose 5-phosphate was present in the assay mixtures.The amount of radioactivity found in CO2, was 6.8-times higher when extracts of methanol-grown Pseudomona C were incubated for a short period of time with [1-14C]glucose 6-phosphate than with [U-14C]glucose 6-phosphate.These data, and the presence of high specific activities of hexulose phosphate synthase, phosphoglucoisomerase, glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase indicate that in methanol-grown Pseudomonas C, formaldehyde carbon is oxidized to CO2 both via a cyclic pathway which includes the enzymes mentioned and via formate as an oxidation intermediate, with the former predominant. 相似文献
9.
Malate saturation isotherms for the NAD+ malic enzyme determined at widely differing, but saturating, concentrations (8, 80, 160 mm) of magnesium show the same response to malate concentration only when velocity is plotted against the concentration of free malate2?. This identification of the ionized malic acid as the true substrate for this enzyme, together with the observation that the complex of Mg with malate has no influence on the reaction rate even at very high concentrations, indicates that the metal ion activator of the enzyme must also bind in the ionized form. A kinetic analysis shows that, with respect to magnesium and malate, the malic enzyme catalyzes a rapid equilibrium reaction of the intersecting type. Either Mg2+ or malate2? may bind first but the fact that the Km's for both Mg2+ and malate2? are smaller than the respective Ki's suggests that, when either metal ion or malate is present on the enzyme, the other is bound more tightly than when it binds to the free enzyme. This demonstration of the nature of the true substrates for this enzyme has implications for studies of the NAD+ malic enzyme in which conditions influencing the amount of free magnesium and malate, e.g., changes in pH, addition of weak acid effectors etc., are involved. 相似文献
10.
The fungicidal mechanism of chlorine dioxide on Saccharomyces cerevisiae was investigated. During S. cerevisiae inactivation by ClO2, protein, DNA, and ion leakage, enzyme activity, genomic DNA structure, and cell ultrastructure were examined. Protein and DNA leakages were not observed, while ion leakages of K+, Ca2+, and Mg2+ were detected and were related to the inactivation rate. The glucose-6-phosphate dehydrogenase, citrate synthase, and phosphofructokinase activities were inhibited and were also correlated with the inactivation rate. Genomic DNA structure was not damaged except for an extremely high ClO2 concentration (100 mg L-1). Electron micrographs showed that cell surface damage was pronounced and disruption in inner cell components was also apparent. The ion leakage, the inhibition of key enzyme activities of metabolic pathway, and the alteration of cell structure were critical events in S. cerevisiae inactivation by ClO2. 相似文献
11.
Structure-activity studies of enzyme substrates using model interaction calculations 总被引:1,自引:0,他引:1
The hydrolysis rates in a series of substituted N-acetylaminoacid methylesters with acylase I depend apparently on the binding strengths of the side chains to the enzyme surface. Using a truncated model of a possible enzyme binding site we calculate the binding energies for a series of straight chain substituents. Our calculations reveal a very good correlation between the calculated energies and the hydrolysis rates, indicating the value of our receptor model. We feel this to be another example of the validity of our receptor mapping using model interaction energy calculations based on the monopoles-bond polarizabilities method. 相似文献
12.
Chorismatase is an important enzyme involved in Shikimate pathway, which catalyzes the conversion of chorismate into pyruvate and (dihydro)‐benzoic acid derivatives. According to the outcomes of catalytic reactions, chorismatases can be divided into three subfamilies: CH‐Fkbo, CH‐Hyg5 and CH‐XanB2. Recently, the crystal structures of CH‐Fkbo and CH‐Hyg5 from Streptomyces hygroscopicus have been successfully obtained, allowing us to perform QM/MM calculations to explore the reaction details. Our calculation results support the proposal that CH‐Fkbo and CH‐Hyg5 employ different catalytic mechanisms and gave the mechanistic details. Fkbo follows a typical hydrolytic mechanism, which contains three consecutive steps, including the protonation step of the methylene group of substrate, the nucleophilic attack of the resulted carbocation by activated water and cleavage of C2′‐O8 bond of tetrahedral intermediate (hemiketal). The protonation of methylene group and the C2′‐O8 cleavage correspond to similar energy barriers (26.5 and 24.8 kcal/mol), suggesting both steps to be rate‐limiting. Whereas Hyg5 employs an intramolecular mechanism, in which the oxygen from C4 migrates to C3 via an arene oxide intermediate. The first step of Hyg5, which corresponds to the concerted protonation of methylene group and the cleavage of C3‐O8, is calculated to be rate‐limiting with an energy barrier of 26.3 kcal/mol. The nonconserved active site residue G240Hyg5 (or A244Fkb°) is suggested to be responsible for leading to different reaction mechanism in CH‐Fkbo and CH‐Hyg5. During the catalytic reaction, residue C327 plays an important role in directing the product selectivity in Hyg5 enzyme. Proteins 2017; 85:1146–1158. © 2017 Wiley Periodicals, Inc. 相似文献
13.
Ramesh N. Patel Ching T. Hou P. Derelanko 《Archives of biochemistry and biophysics》1983,221(1):135-142
An NAD+-linked, reduced glutathione-dependent formaldehyde dehydrogenase was purified to homogeneity from soluble extracts of methanol-grown yeast, Pichia sp. Formaldehyde and methylglyoxal are oxidized in the presence of NAD+ as an electron acceptor. NADP+ could not replace NAD+. Other straight chain aldehydes (C2–C6 tested), branched-chain aldehydes (e.g., isobutyaldehyde), aromatic aldehydes (e.g., salicylal-dehyde, benzaldehyde), glutyraldehyde, glyceraldehyde, glycoaldehyde, and glyoxal-dehyde tested were not oxidized by the purified formaldehyde dehydrogenase. The product of formaldehyde oxidation by purified enzyme was demonstrated to be S-for-mylglutathione by measuring the absorption at 240 nm due to the formation of thioester of formaldehyde and reduced glutathione. The Km values for NAD+, formaldehyde, and reduced glutathione were 0.12, 0.31, and 0.16 mm, respectively, for the forward reaction at pH 8.0. The purified formaldehyde dehydrogenase also catalyzed the reduction of S-formylglutathione in the presence of NADH. Formate was not reduced by the purified enzyme. The Km values for S-formylglutathione and NADH were 0.60 and 0.25 mm, respectively, for the reverse reaction at pH 6.0. Formaldehyde dehydrogenase has a molecular weight of 84,000 as determined by gel filtration and subunit molecular weight of 41,000 as determined by sodium dodecyl sulfate-gel electrophoresis. S-Formylglutathione, a product of formaldehyde oxidation, was oxidized by the partially purified formate dehydrogenase from Pichia sp. Formate dehydrogenase has a higher affinity toward S-formylglutathione (Km value 1.8 mm) than toward formate (Km value 25 mm). Antiserum prepared against the purified formaldehyde dehydrogenase from Pichia sp. NRRL-Y-11328 forms strong precipitin bands with isofunctional enzymes from methanol-grown Pichia pastoris NRRL-Y-7556 and Torulopsis candida Y-11419 and weak precipitin bands with Hansenula polymorpha NRRL-Y-2214. No cross-reaction was observed with isofunctional enzyme derived from methanol-grown Kloeckera sp. 相似文献
14.
Dr. T. H. Özdamar Dr. S. Takaç Dr. G. Çalik R. Ballica 《Bioprocess and biosystems engineering》1989,4(6):249-256
Kinetics of the separation of L-glutamic acid (GLU) by ion exchange has been studied with strongly acidic H+-type cation exchange resin Amberlite IR-122. Since glutamic acid is a trivalent ampholyte and dissociates according to three equilibrium reactions, separation of G+ ions by a cation exchange process is accompanied by the dominant reversible reaction, i.e. G++H+ ? G0. Accompanying reversible reaction has an effect on the ion exchange rate, and decreases the performance of the process comparing with the ideal case that the exchanging ions retain their identity. The analysis was performed first with the ion exchange column, DIC (L/D=0.52); and then with the ion exchange column, IC (L/D=10.9). The data were collected with model glutamic acid solutions for both DIC and IC columns/reactors. IC experimental results were then compared with that of DIC and the effect of scale up on ion exchange process was investigated. The experimental results have provided an adequate basis for the design calculations, and the design parameters were determined. Rate coefficients for the liquid phase mass transfer controlled cation exchange process were calculated and interrelated with a plot of j Mfactor versus Reynolds number. 相似文献
15.
Ina Bisha Alex Rodriguez Alessandro Laio Alessandra Magistrato 《PLoS computational biology》2014,10(12)
Sodium-Galactose Transporter (SGLT) is a secondary active symporter which accumulates sugars into cells by using the electrochemical gradient of Na+ across the membrane. Previous computational studies provided insights into the release process of the two ligands (galactose and sodium ion) into the cytoplasm from the inward-facing conformation of Vibrio parahaemolyticus sodium/galactose transporter (vSGLT). Several aspects of the transport mechanism of this symporter remain to be clarified: (i) a detailed kinetic and thermodynamic characterization of the exit path of the two ligands is still lacking; (ii) contradictory conclusions have been drawn concerning the gating role of Y263; (iii) the role of Na+ in modulating the release path of galactose is not clear. In this work, we use bias-exchange metadynamics simulations to characterize the free energy profile of the galactose and Na+ release processes toward the intracellular side. Surprisingly, we find that the exit of Na+ and galactose is non-concerted as the cooperativity between the two ligands is associated to a transition that is not rate limiting. The dissociation barriers are of the order of 11–12 kcal/mol for both the ion and the substrate, in line with kinetic information concerning this type of transporters. On the basis of these results we propose a branched six-state alternating access mechanism, which may be shared also by other members of the LeuT-fold transporters. 相似文献
16.
17.
A Mn2+-activated phosphohistone phosphatase has been isolated from canine heart. The s20, w for the enzyme is 3.8. Using this value and the value for Stokes radius (39 Å), the molecular weight for the enzyme was calculated to be 61,000. The enzyme is inactive in the absence of divalent cations, among which Mn2+ is the most effective activator. Co2+ and Mg2+ are less effective than is Mn2+. Zn2+, Fe2+, and Cu2+ are inhibitory. The enzyme has a pH optimum between 7 and 7.5 and has an apparent Km for phosphohistone and Mn2+ of about 17 μm and 0.5 mm, respectively. The enzyme is inhibited by nucleoside triphosphate, ADP, AMP, phosphate, and pyrophosphate, but is not affected by cyclic AMP or cyclic GMP. The dephosphorylation of phosphohistone is stimulated by salts. Kinetic studies reveal that KCl and other salts greatly affect both the rate of hydrolysis and the Km for either Mn2+ or phosphohistone by interacting with the substrate. The data suggest that modification at substrate level is an important regulatory mechanism for the enzyme. The enzyme preparation also dephosphorylates phosphorylase a and phosphocasein. Evidence suggests that one enzyme possesses both phosphohistone and phosphorylase phosphatase activities and that a different enzyme catalyzes the Mg2+- and Mn2+-activated dephosphorylation of phosphocasein. 相似文献
18.
Halomethane:Bisulfide/Halide Ion Methyltransferase, an Unusual Corrinoid Enzyme of Environmental Significance Isolated from an Aerobic Methylotroph Using Chloromethane as the Sole Carbon Source 总被引:4,自引:4,他引:0
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Catherine Coulter John T. G. Hamilton W. Colin McRoberts Leonid Kulakov Michael J. Larkin David B. Harper 《Applied microbiology》1999,65(10):4301-4312
A novel dehalogenating/transhalogenating enzyme, halomethane:bisulfide/halide ion methyltransferase, has been isolated from the facultatively methylotrophic bacterium strain CC495, which uses chloromethane (CH3Cl) as the sole carbon source. Purification of the enzyme to homogeneity was achieved in high yield by anion-exchange chromatography and gel filtration. The methyltransferase was composed of a 67-kDa protein with a corrinoid-bound cobalt atom. The purified enzyme was inactive but was activated by preincubation with 5 mM dithiothreitol and 0.5 mM CH3Cl; then it catalyzed methyl transfer from CH3Cl, CH3Br, or CH3I to the following acceptor ions (in order of decreasing efficacy): I−, HS−, Cl−, Br−, NO2−, CN−, and SCN−. Spectral analysis indicated that cobalt in the native enzyme existed as cob(II)alamin, which upon activation was reduced to the cob(I)alamin state and then was oxidized to methyl cob(III)alamin. During catalysis, the enzyme shuttles between the methyl cob(III)alamin and cob(I)alamin states, being alternately demethylated by the acceptor ion and remethylated by halomethane. Mechanistically the methyltransferase shows features in common with cobalamin-dependent methionine synthase from Escherichia coli. However, the failure of specific inhibitors of methionine synthase such as propyl iodide, N2O, and Hg2+ to affect the methyltransferase suggests significant differences. During CH3Cl degradation by strain CC495, the physiological acceptor ion for the enzyme is probably HS−, a hypothesis supported by the detection in cell extracts of methanethiol oxidase and formaldehyde dehydrogenase activities which provide a metabolic route to formate. 16S rRNA sequence analysis indicated that strain CC495 clusters with Rhizobium spp. in the alpha subdivision of the Proteobacteria and is closely related to strain IMB-1, a recently isolated CH3Br-degrading bacterium (T. L. Connell Hancock, A. M. Costello, M. E. Lidstrom, and R. S. Oremland, Appl. Environ. Microbiol. 64:2899–2905, 1998). The presence of this methyltransferase in bacterial populations in soil and sediments, if widespread, has important environmental implications. 相似文献
19.
The kinetics of acetylcholinesterase alkylation with N,N-dimethyl-2-phenylaziridinium ion, the anionic-site-directed affinity label, has been investigated in the presence of alkylboronic acids, which are known as the esteratic-site-directed reversible inhibitors of the enzyme. The ternary complex of the enzyme, the aziridinium ion and alkylboronic acid, are formed in this reaction. In the case of propylboronic acid, for which the complete kinetic analysis of the acceleration effect has been carried out, the 85-fold increase in the rate of the enzyme alkylation reaction has been found. This acceleration effect was connected with the alkylation step, whereas the non-covalent binding of the aziridinium ion in the enzyme active centre was even hindered by the alkylboronic acid. The possible mechanism of this kinetic acceleration phenomenon is discussed with special reference to the kinetic data for the spontaneous solvolysis reaction of the aziridinium ion in water and organic solvents. 相似文献
20.
《Biochimica et Biophysica Acta - Proteins and Proteomics》2018,1866(7):759-766
The l-α-Asp residues in peptides or proteins are prone to undergo nonenzymatic reactions to form l-β-Asp, d-α-Asp, and d-β-Asp residues via a succinimide five-membered ring intermediate. From these three types of isomerized aspartic acid residues, particularly d-β-Asp has been widely detected in aging tissue. In this study, we computationally investigated the cyclization of α- and β-Asp residues to form succinimide with dihydrogen phosphate ion as a catalyst (H2PO4−). We performed the study using B3LYP/6-31 + G(d,p) density functional theory calculations. The comparison of the activation barriers of both residues is discussed. All the calculations were performed using model compounds in which an α/β-Asp-Gly sequence is capped with acetyl and methylamino groups on the N- and C-termini, respectively. Moreover, H2PO4− catalyzes all the steps of the succinimide formation (cyclization-dehydration) acting as a proton-relay mediator. The calculated activation energy barriers for succinimide formation of α- and β-Asp residues are 26.9 and 26.0 kcal mol− 1, respectively. Although it was experimentally confirmed that β-Asp has higher stability than α-Asp, there was no clear difference between the activation barriers. Therefore, the higher stability of β-Asp residue than α-Asp residue may be caused by an entropic effect associated with the succinimide formation. 相似文献