Nonenzymic Model Research Articles

Formation of Strecker Aldehydes from Polyphenol-Derived Quinones and α-Amino Acids in a Nonenzymic Model System

Fruits and vegetables contain naturally occurring polyphenolic compounds that can undergo enzyme-catalyzed oxidation during food preparation. Many of these compounds contain catechol (1,2-dihydroxybenzene) moieties that may be transformed into o-quinone derivatives by polyphenoloxidases and molecular oxygen. Secondary reactions of the o-quinones include the Strecker degradation of ambient amino acids to form flavor-important volatile aldehydes. The purpose of this work was to investigate the mechanism of the polyphenol/o-quinone/Strecker degradation sequence in a nonenzymic model system. By using ferricyanide ion as the oxidant in pH 7.17 phosphate buffer at 22 degrees C, caffeic acid, chlorogenic acid, (+) catechin, and (-) epicatechin were caused to react with methionine and phenylalanine to produce Strecker aldehydes methional and phenylacetaldehyde in 0.032-0.42% molar yields (0.7-10 ppm in reaction mixtures). Also, by employing l-proline methyl ester in a reaction with 4-methylcatechol, a key reaction intermediate, 4-(2'-carbomethoxy-1'-pyrrolidinyl)-5-methyl-1,2-benzoquinone (7), was isolated and tentatively identified.

Degradation of β-lactam antibiotics in the presence of Zn 2+ and 2-mino-2-hydroxymethylpropane-1,3-diol (Tris). A hypothetical non-enzymic model of β-lactamases

The system composed of 2-amino-2-hydroxymethylpropane-1,3-diol (Tris) and Zn 2+ catalyses the degradation of cephalosporins. The β-lactam opening fits to a first-order process, with a constant directly proportional to the zinc ion concentration. The pH and Tris concentration dependency displayed by the first-order constant, as well as the nature of the degradation products point to a mechanism that can be considered as an extension of that proposed for the benzylpenicillin degradation. The mechanism proposed here, and the values of the kinetic constants calculated, as compared with those of β-lactamases, lead to the conclusion that the Tris-Zn 2+ system simulates the catalytic action of the serine β-lactamases rather than the action of the Zn 2+-dependent type of enzymes.

Conversion of 19-oxo[2β-2H]androgens into oestrogens by human placental aromatase. An unexpected stereochemical outcome

Aromatase is a cytochrome P-450 enzyme that catalyzes the conversion of androgens into oestrogens via sequential oxidations at the 19-methyl group. Despite intensive investigation, the mechanism of the third step, conversion of the 19-aldehydes into oestrogens, has remained unsolved. We have previously found that a pre-enolized 19-al derivative undergoes smooth aromatization in non-enzymic model studies, but the role of enolization by the enzyme in transformations of 19-oxoandrogens has not been previously investigated. The compounds 19-oxo[2 beta-2H]testosterone and 19-oxo[2 beta-2H]androstenedione have now been synthesized. Exposure of either of these compounds to microsomal aromatase, in the absence of NADPH, for an extended period led to no significant 2H loss or epimerization at C-2, leaving open the importance of an active-site base. However, in the presence of NADPH there was an unexpected substrate-dependent difference in the stereoselectivity of H loss at C-2 in the enzyme-induced aromatization of 19-oxo[2 beta-2H]-testosterone versus 19-oxo[2 beta-2H]androstenedione. The aromatization results for 17 beta-ol derivative 19-oxo[2 beta-2H]-testosterone correspond to about 1.2:1 2 beta-H/2 alpha-H loss from unlabelled 19-oxotestosterone. In contrast, aromatization results for 19-oxo[2 beta-2H]androstenedione correspond to at least 11:1 2 beta-H/2 alpha-H loss from unlabelled 19-oxoandrostenedione. This substrate-dependent stereoselectivity implies a direct role for an enzyme active-site base in 2-H removal. Furthermore, these results argue against the proposal that 2 beta-hydroxylation is the obligatory third step in aromatase action.

Metal-catalyzed oxidation of Escherichia coli glutamine synthetase: Correlation of structural and functional changes

Metal-catalyzed oxidation of proteins has been implicated in a variety of biological processes, particularly in the marking of proteins for subsequent proteolytic degradation. The metal-catalyzed oxidation of bacterial glutamine synthetase causes conformational, covalent, and functional alterations in the protein. To understand the structural basis of the functional changes, the time course of oxidative modification of glutamine synthetase was studied utilizing a nonenzymic model oxidation system consisting of ascorbate, oxygen, and iron. The structural modifications induced included: decreased thermal stability; weakening of subunit interactions; decrease in isoelectric point; introduction of carbonyl groups into amino acid side chains; and loss of two histidine residues. These changes did not denature the protein, but instead induced relatively subtle changes. Indeed, even the most extensively modified protein had a sedimentation velocity which was identical to that of the native enzyme. Comparison of the time courses of the structural and functional changes established that: (i) Loss of the metal binding site and of catalytic activity occurred with loss of one histidine per subunit; (ii) increased susceptibility to proteolysis occurred with loss of two histidine residues per subunit. Thus, oxidation at one site suffices to inactivate the enzyme, but two sites must be modified to induce susceptibility to proteolysis. The limited and specific changes induced by metal-catalyzed oxidation are consistent with a site-specific free radical mechanism.

On the mechanism of action of vitamin K. A new nonenzymic model

On the mechanism of action of vitamin K. A new nonenzymic model

The Fenton degradation as a nonenzymic model for microsomal denitrosation of N-nitrosodimethylamine

The microsomal metabolism of the carcinogen N-nitrosodimethylamine (NDMA) was suggested to be initiated by hydrogen atom abstraction to form an alpha-nitrosamino radical, which either oxidizes further to an alpha-hydroxy nitrosamine as the initial product of the activating dealkylation pathway or fragments to the nitric oxide radical and N-methylformaldimine as the first step of the presumably inactivating denitrosation route. To examine the chemistry of the alpha-nitrosamino radical in a nonenzymatic setting, we exposed NDMA to the Fenton reagent, which is known to be capable of abstracting hydrogen atoms from organic species. The products observed were those expected of a denitrosation model. Solutions containing 13 mM [14C]NDMA, 15 mM FeSO4, 15 mM H2O2, and 7.5 mM H2SO4 were kept at 4-10 degrees C for 1 h and then basified to yield methylamine (3.2 +/- 0.5 mM, mean +/- SD, n = 8), formaldehyde (3.1 +/- 0.9 mM), and unreacted nitrosamine (10.2 +/- 0.7 mM) as the only radioactive species detected, with total nitrate/nitrite also being found at a level of 2.8 +/- 0.5 mM. N-Methylformaldiminium ion was identified as an intermediate. The parallels between these results and those seen in the microsomal reaction support the hypothesis that the alpha-nitrosamino radical is a common intermediate in enzymatic denitrosation versus dealkylation of NDMA.(ABSTRACT TRUNCATED AT 250 WORDS)

Aminopropyltransferases: mechanistic studies and the synthesis of specific inhibitors.

We have studied the mechanism of enzyme-catalyzed alkyl transfer reactions by a variety of methods including non-enzymic model reactions, steady-state kinetics, and stereochemistry. In this brief paper, the results of our earlier work on catechol O-methyltransferase (COMT, E.C. 2.1.1.6) and more recent work on putrescine aminopropyltransferase (PAPT, E.C. 2.5.1.16, sometimes referred to as spermidine synthase) will be reviewed. These investigations have resulted in the demonstration that each of these alkyltransferases catalyze a reaction which proceeds via a ternary complex involving the enzyme and both nucleophilic and electrophilic substrates. In addition, the demonstration of general base catalysis in related model reactions, provides chemical precedent for the involvement of this type of catalysis by basic residues of the enzyme. The mechanistic conclusions have allowed us to design and synthesize several mechanism-based inhibitors which exhibit great potency and specificity for individual alkyltransferases1.

The mechamism of action of vitamin B 12

A novel rearrangement reaction is introduced as a model for the rearrangement of methylitaconic acid (III) to α-methyleneglutaric acid (IV), one of three enzyme catalyzed, coenzyme B 12-dependent, carbonskeleton rearrangements whose mechanism has been a source of puzzlement for many years. The key feature of the new model is the direct attachment of the substrate, methylitaconic acid, to the cobalt atom of vitamin B 12 This was accomplished by reacting butadiene-2,3-decarboxylic acid with hydrobromic acid generating bromomethylitaconic acid (VIII). Use of two moles of hydrobromic acid yielded bis-2,3-(bromomethyl)succinic acid (IX). Reaction of the monobromide VIII with vitamin B 12s did not yield the desired carbon-cobalt bonded adduct. Instead, the lactone ηa- methylene-γbutyrolactone-β-carboxylic acid (X) was formed. Accordingly, the ester, dimethyl bromomethylitaconate (XIa), was reacted with vitamin B 12s and yielded the carbon-cobalt bonded adduct XIIa. Bis-trimethylsilyl bromomethylitaconate did not yield an adduct when reacted with vitamin B 12s, but bis-tetrahydropyranyl bromomethylitaconate (XIb) did yield the adduct XIIb. The ester cobalamin XIIb undergoes spontaneous decomposition at room temperature, in aqueous solution, at pH 8 and in the dark - biochemically ideal circumstances - yielding a mixture of butadiene-2,3-decarboxylic acid (VII), methylitaconic acid (III) and α- methyleneglutaric acid (IV). The presence of the latter indicates that a skeletal change has taken place in a way which mimics the enzymatic reaction. This is the first non-enzymic model in this carbon-skeleton rearrangement series. The methyl ester cobalamin XIIa was stable in the dark but did decompose on irradiation with a sunlamp to butadiene-2,3decarboxylic acid (VII) and methylitaconic acid (III). No α-methyleneglutaric acid IV was observed in the latter reaction. Authentic methylitaconic acid (III) was prepared by alkylation of triethyl prop-2-ene-l,l,2-tricarboxylate (XIII) with methyl iodide followed by hydrolysis and decarboxylation. The lactone X and lactone α-methyl-γ-butyrolactone-β-carboxylic acid (XVI) were prepared by condensing the triester XIII with formaldehyde, hydrolyzing the lactone diester XV to the lactone X and hydrogenating to the saturated lactone XVI.

A nonenzymic model for the coenzyme B 12-dependent isomerization of methylmalonyl-SCoA to suiccinyl-SCoA

Dimethyl bromomethylmalonate (IV) reacts with vitamin B 12s in aqueous solution yielding a relatively unstable carbon-cobalt bonded adduct V, which shows visible spectra in good accord with expectation. The adduct V was allowed to decompose in water, in the dark, at room temperature and at physiological pH. Three products: succinic acid (VI), methylmalonic acid (VIII) and malonic acid (VII) were formed in 3, 18, and 13% yields respectively. Isolation of the succinic acid rearrangement product provides support for the intermediacy of the carbon-cobalt bond in the coenzyme B 12 dependent enzymic carbon-skeleton rearrangement of methylmalonyl-SCoA to succinyl-SCoA.

Inactivation of D-serine dehydratase by alkylamines via a transimination of enzyme-linked cofactor.

The time-dependent inactivation of D-serine dehydratase by alkylamines was characterized. Evidence is presented indicating that inactivation proceeds via a transimination reaction analogous to the first step in the catalytic pathway. The product of alkylamine attack, an alkylamine pyridoxal 5'-phosphate Schiff base, readily dissociated from D-serine dehydratase to produce inactive apoenzyme. Reaction with alkylamines was shown to be a convenient way of producing apoenzyme which can be reconstituted with pyridoxal 5'-phosphate to fully active holoenzyme. Amino acids such as glycine and alanine, unlike alkylamines, did not resolve D-serine dehydratase but were competitive inhibitors of the enzyme. The inability of amino acids to resolve D-serine dehydratase from its cofactor was attributed to a failure of the amino acid-cofactor Schiff base to dissociate from the enzyme. Transimination of D-serine dehydratase with its substrate D-serine was at least 3.5 X 10(5) times faster than a nonenzymic model transimination reaction and more than 70 million times faster than the reaction of the enzyme with 2-hydroxyethylamine, indicating that the carboxyl group of the substrate is an important structural determinant for catalysis of the transimination step in the catalytic pathway. An analysis of the inhibitory effect of potassium ion on the rate and extent of inactivation of D-serine dehydratase by alkylamines indicated that K+ binding increased the affinity of the enzyme for its cofactor by at least 13-fold.

Electrochemical energetics of biochemical hydroxylations and of their non-enzymic models

The biochemical and physiological importance of hydroxylations, which permit the primary oxidation of hydrogenerated carbon atoms using oxygen partly reduced by bielectronic reducing agents without mediation of the electron transfer chain, is reviewed. The energetic and mechanistic data obtained from electrochemical studies of oxygen reduction permit to discuss the mechanisms of these hydroxylations. Artificial oxidations at room temperature in aqueous or organic solvents by so-called activated oxygen molecules or Fenton reagents involving hydrogen peroxide must be considered as non-enzymic analogs of biochemical hydroxylations. The energetics and kinetics of the reduction of oxygen to hydrogen peroxide by hydroquinone, followed by degradative oxidation of the produced quinone, representing a non-enzymic model of the biochemical oxidation of homogentisate, was experimentally studied and discussed from available data related to the energetics of redox couples involving hydrogen peroxide and superoxide ion radical. The field of application in chemistry and in physiology and medicine of such non-enzymic analogs of biochemical hydroxylations performed either chemically or electrochemically, is briefly reviewed.

Nonenzymic model of the dioldehydratase reaction. Initiation of conversion of ethylene glycol into acetaldehyde on anaerobic photolysis of organocobalamins in the presence of SH-compounds

Anaerobic photolysis of adenosyl- and methyl-cobalamins in an aqueous KCl–HCl buffer (pH 2·0) in the presence of SH-compounds (e.g. dihydrolipoic acid amide) causes conversion of ethylene glycol into acetaldehyde, and provides a model for the dioldehydratase enzymic reaction.

Letter: A nonenzymic model intermediate for the coenzyme B 12 dependent isomerization of methymalonyl coenzyme A to succinyl coenzyme A.

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTA nonenzymic model intermediate for the coenzyme B12 dependent isomerization of methylmalonyl coenzyme A to succinyl coenzyme APaul Dowd and Moritz ShapiroCite this: J. Am. Chem. Soc. 1976, 98, 12, 3724–3725Publication Date (Print):June 1, 1976Publication History Published online1 May 2002Published inissue 1 June 1976https://doi.org/10.1021/ja00428a065RIGHTS & PERMISSIONSArticle Views85Altmetric-Citations28LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (317 KB) Get e-Alerts Get e-Alerts

OXYGENATION OF 3-SUBSTITUTED INDOLES CATALYZED BY Co(II)-SCHIFF′S BASE COMPLEXES. A MODEL CATALYTIC OXYGENATION FOR TRYPTOPHAN 2,3-DIOXYGENASE

Abstract Bis(salicylidene)ethylenediaminatocobalt(II) catalyzes the oxygenation of 3-substituted indoles related to tryptophan in methanol giving rise to oxidative cleavage of the heterocyclic ring of the indoles selectively to give the corresponding o-formylaminoacetophenone derivatives. This provides a nonenzymic model for the reaction of tryptophan 2,3-dioxygenase. Oxidizability of the substrates is correlated with their donor ability.

A model catalytic oxygenation for the reaction of quercetinase

Bis(salicylidene)ethylenediaminatocobalt(II) catalyses the oxygenation of 3-hydroxyflavones in dimethylformamide giving rise to oxidative cleavage of the heterocyclic ring of the flavones to give the corresponding depsides in excellent yield; this provides a nonenzymic model for the reaction of quercetinase.

Base-catalysed autoxidation of 3,4′-dihydroxyflavone

Autoxidation of 3,4′-dihydroxyflavone in dimethylformamide in the presence of potassium t-butoxide results in oxidative cleavage of the hetrocyclic ring to give 2-(4-hydroxybenzoyloxy) benzoic acid and carbon monoxide in excellent yields; this provides a nonenzymic model for the reaction of quercetinas.

Synthesis of l-thyroxine from 3,5-diiodo- l-tyrosine by nonenzymic transamination

A nonenzymic model for the biosynthesis of thyroxine from diiodotyrosine is described. In this reaction, l-thyroxine is formed from 3,5-diiodo- l-tyrosine at neutral pH and at room temperature in the presence of sodium glyoxylate, cupric acetate, and oxygen. The reaction proceeds rapidly through transamination via a metal chelate of the Schiff base of diiodotyrosine and glyoxylic acid, followed by oxidative coupling of the 4-hydroxy-3,5-diiodophenylpyruvic acid thus formed, with diiodotyrosine.

Reactivating effect of some naturally occurring dithiol compounds on threonine deaminase

The biodegradative threonine deaminases have been purified to homogeneity and crystallized from the extracts of Escherichia coli (E. coli) and Clostridium tetranomorphum, respectively. It has been demonstrated that deaminases are composed of four identical subunits, each monomer containing one mole of pyridoxal phosphate. The deamination reaction mechanism studied using the E. coli enzyme is compatible with the mechanism derived from the pyridoxal-catalyzed α,β-elimination reaction in the non-enzymic model system, although direct evidence for the participation of pyridoxal phosphate in the enzyme reaction is lacking. Spectral and circular dichroic changes of the enzyme-bound pyridoxal phosphate after addition of the substrate, L-threonine, were postulated to represent the formation of a complex between pyridoxal phosphate and the dehydrated reaction intermediate. Optical studies indicate that optical changes occur prior to the β-elimination reaction. Kinetic analyses have indicated that the enzyme exists as an inactive form under the conditions of these optical experiments. These phenomena could be interpreted as indicating that the conversion of this inactive enzyme into an active form occurs after addition of L-threonine. These optical changes are observed during the reaction and this conversion step appears to be the rate-limiting step of the overall reaction under the conditions employed.

Deuterium migration during the acid-catalyzed dehydration of 6-deuterio-5,6-dihydroxy-3-chloro-1,3-cyclohexadi-ene, a nonenzymic model for the NIH shift

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTDeuterium migration during the acid-catalyzed dehydration of 6-deuterio-5,6-dihydroxy-3-chloro-1,3-cyclohexadi-ene, a nonenzymic model for the NIH shiftDonald M. Jerina, John W. Daly, and Bernhard. WitkopCite this: J. Am. Chem. Soc. 1967, 89, 21, 5488–5489Publication Date (Print):October 1, 1967Publication History Published online1 May 2002Published inissue 1 October 1967https://doi.org/10.1021/ja00997a053RIGHTS & PERMISSIONSArticle Views55Altmetric-Citations31LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (236 KB) Get e-Alerts Get e-Alerts