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Upendra K. Pandit - One of the best experts on this subject based on the ideXlab platform.
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Alkyl transfer from quaternary ammonium salts to cobalt (I): Model for the cobalamin-dependent methionine Synthase Reaction
Tetrahedron, 1994Co-Authors: Ellen Hilhorst, Atef S. Iskander, Tjoe B.r.a. Chen, Upendra K. PanditAbstract:Abstract The Reaction of cobaloxime(I) with diverse quaternary ammonium salts leads, in general, to a group transfer from nitrogen to cobalt. The behaviour of the salts in these transalkylations is consistent with an SN2 mechanism, involving Co(I) as a nucleophile. In a model study of the cobalamin-dependent methionine Synthase Reaction, 5-13CH3-methyl labelled 5,5,6,7-tetramethyl-5,6.7,8-tetrahydropteridinium salt(23) -a model of the natural coenzyme 5-CH3H4-folate (1)- was allowed to react with cobaloxime(I) and cobalamin(I). In each case the formation of the methyl transfer product, namely, methylcobaloxime and mothylcobalamin, respectively, was shown by 13C-NMR spectroscopy.
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of Quaternary Ammonium Salts by Thiolate Anions: A Model of the Cobalamin-independent Methionine Synthase Reaction.
1994Co-Authors: Ellen Hilhorstl, Atef S. Wander, Upendra K. PanditAbstract:The Reactions of thiolate ions derived from thiophenol and homocysteioe with substituted quatemary ammoniom salts result in alkyl tmnsfer from nitrogen to sulfur. A radical mechanism for this tmnsakylation, accounts for the reactivity pattern of the substrate salts. In a model study of the cobalamin-independent methionine Synthase Reaction, 5,5.6,7-tetramerhyl-5.6;7,8-tetrahydro- ptcridiniumsalt(25),whichcanbcconsidaedasamodelf~thenaMal UxxEymc 5-CH$-I4-folate (1). was allowed to react with the thiilate of homocysteiae, whereupon the won of m&ion& was observed in good yield. 'These results suggest that in the enzymatic prcnxss the N(S)-CH3 bond may be activated for the methyl transfer step, by uxndination of the N(5) with an electmphde or a proton at the active site.
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Dealkylation of quaternary ammonium salts by thiolate anions: A model of the cobalamin-independent methionine Synthase Reaction.
Tetrahedron, 1994Co-Authors: Ellen Hilhorst, Atef S. Iskander, Tjoe B.r.a. Chen, Upendra K. PanditAbstract:Abstract The Reactions of thiolate ions derived from thiophenol and homocysteine with substituted quaternary ammonium salts result in alkyl transfer from nitrogen to sulfur. A radical mechanism for this transalkylation, accounts for the reactivity pattern of the substrate salts. In a model study of the cobalamin-independent methionine Synthase Reaction, 5,5,6,7-tetramethyl-5,6,7,8-tetrahydropteridinium salt (25), which can be considered as a model for the natural coenzyme 5-CH3H4-folate (1), was allowed to react with the thiolate of homocysteine, whereupon the formation of methionine was observed in good yield. These results suggest that in the enzymatic process the N(5)-CH3 bond may be activated for the methyl transfer step, by coordination of the N(5) with an electrophile or a proton at the active site.
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Model studies of the cobalamin-dependent methionine Synthase Reaction
Tetrahedron Letters, 1993Co-Authors: Ellen Hilhorst, Atef S. Iskander, Tjoe B.r.a. Chen, Upendra K. PanditAbstract:Abstract The cobalt atom of cobaloxime(I) and cobalamin(I) is methylated by a 5-methylpterinium salt acting as a model of the activated 5-methyltetrahydrofolate cofactor.
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A model of the cobalamin-independent methionine Synthase Reaction
Journal of the Chemical Society Chemical Communications, 1993Co-Authors: Ellen Hilhorst, Tjoe B.r.a. Chen, Upendra K. PanditAbstract:Homocysteine is converted to methionine via a nonenzymatic methyl transfer from a 5-methyitetrahydrofolate model bearing a positive charge at N(5).
Chris Abell - One of the best experts on this subject based on the ideXlab platform.
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A Secondary β Deuterium Kinetic Isotope Effect in the Chorismate Synthase Reaction
Bioorganic chemistry, 2000Co-Authors: Stephen Bornemann, Maria-elena Theoclitou, Martin Brune, Martin R. Webb, Roger N. F. Thorneley, Chris AbellAbstract:Abstract Chorismate Synthase (EC 4.6.1.4) is the shikimate pathway enzyme that catalyzes the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate. The enzyme Reaction is unusual because it involves a trans -1,4 elimination of the C-3 phosphate and the C-6 proR hydrogen and it has an absolute requirement for reduced flavin. Several mechanisms have been proposed to account for the cofactor requirement and stereochemistry of the Reaction, including a radical mechanism. This paper describes the synthesis of [4- 2 H]EPSP and the observation of kinetic isotope effects using this substrate with both Neurospora crassa and Escherichia coli chorismate Synthases. The magnitude of the effects were D ( V ) = 1.08 ± 0.01 for the N. crassa enzyme and 1.10 ± 0.02 on phosphate release under single-turnover conditions for the E. coli enzyme. The effects are best rationalised as substantial secondary β isotope effects. It is most likely that the C(3)-O bond is cleaved first in a nonconcerted E1 or radical Reaction mechanism. Although this study alone cannot rule out a concerted E2-type mechanism, the C(3)-O bond would have to be substantially more broken than the proR C(6)-H bond in a transition state of such a mechanism. Importantly, although the E. coli and N. crassa enzymes have different rate limiting steps, their catalytic mechanisms are most likely to be chemically identical.
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AN ELECTROCHEMICAL STUDY TO MODEL THE CHORISMATE Synthase Reaction
Bioorganic & Medicinal Chemistry Letters, 1996Co-Authors: Maria-elena Theoclitou, Peter J. Duggan, Chris AbellAbstract:Abstract The electrochemical reduction of 3-phosphate-cyclohex-1-ene-1-carboxylic acid 13 has been studied as a model for the chorismate Synthase Reaction. An electrochemical Reaction occurs only when 13 is fully protonated. However the Reaction results in loss of the carboxyl group rather than the phosphate.
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Observation of a secondary tritium isotope effect in the chorismate Synthase Reaction.
Biochemistry, 1995Co-Authors: Sundaram Balasubramanian, John R. Coggins, Chris AbellAbstract:Chorismate Synthase, the seventh enzyme on the shikimate pathway, catalyzes the formation of chorismate from 5-enolpyruvylshikimate 3-phosphate (EPSP). This Reaction involves the loss of phosphate from C(3) and hydrogen from the C(6) pro-R position of EPSP. In order to probe the mechanism of this Reaction, [3-(3)H, 14C]EPSP has been synthesized and a secondary V/K tritium kinetic isotope measured for the Reaction catalyzed by Neurospora crassa chorismate Synthase. A small but significant value of kH/kT = 1.047 +/- 0.012 was observed. The Reaction is also shown to be effectively irreversible. Previous experiments have measured a primary deuterium isotope effect on V/K at C(6) [Balasubramanian, S., Abell, C., & Coggins, J. R. (1990) J. Am. Chem. Soc. 112, 8581-8583], and there is additionally evidence in support of a flavin intermediate in the mechanism [Ramjee, M. N., Coggins, J. R., Hawkes, T. R., Lowe, D. J., & Thorneley, R. N. F. (1991) J. Am. Chem. Soc. 113, 8566-8567]. In the light of these observations the Reaction mechanism probably involves cleavage of the C(6)-H and C(3)-O bonds in distinct but partially rate determining steps.
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The synthesis of (6R)-[6-2H]- and (6S)-[6-2H]5-enolpyruvylshikimate-3-phosphate
Tetrahedron Letters, 1991Co-Authors: Shankar Balasubramanian, Chris AbellAbstract:Abstract (6R)-[6-2H]- and (6S)-[6-2H]-enolpyruvylshikimate-3-phosphate have been synthesised for the kinetic analysis of the chorismate Synthase Reaction. The synthesis uses purified shikimate kinase and EPSP Synthase for enantiospecific biotransformations and also clarifies the stereochemical course of the Diels Alder Reaction in the synthesis of shikimic acid.
Po-huang Liang - One of the best experts on this subject based on the ideXlab platform.
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product distribution and pre steady state kinetic analysis of escherichia coli undecaprenyl pyrophosphate Synthase Reaction
Biochemistry, 2000Co-Authors: Jianjung Pan, Sheantai Chiou, Po-huang LiangAbstract:Undecaprenyl pyrophosphate Synthase (UPPs) catalyzes the condensation of eight molecules of isopentenyl pyrophosphate (IPP) with farnesyl pyrophosphate (FPP) to generate C55 undecaprenyl pyrophosphate. We investigated the kinetics and mechanism of this Reaction pathway using Escherichia coli UPPs. With a variety of different ratios of enzyme to substrate and FPP to IPP in the presence or absence of Triton, different product distributions were found. In the presence of excess FPP, the intermediates (C25−C50) accumulated. Under a condition with enzyme and FPP in excess of IPP, instead of C20-geranylgeranyl pyrophosphate, C20, C25, and C30 were the major products. The UPPs steady-state kcat value (2.5 s-1) in the presence of 0.1% Triton was 190-fold larger than in the absence of Triton (0.013 s-1). The kcat value matched the rate constant of each IPP condensation obtained from the enzyme single-turnover experiments. This suggested that the IPP condensation rather than product release was the rate-limiting st...
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Pre-steady-state kinetic analysis of the trichodiene Synthase Reaction pathway.
Biochemistry, 1997Co-Authors: David E. Cane, Hsien Tai Chiu, Po-huang Liang, Karen S AndersonAbstract:The pre-steady-state kinetics of the trichodiene Synthase Reaction were investigated by rapid chemical quench methods. The single-turnover rate was found to be 3.5−3.8 s-1, a rate 40 times faster than the steady-state catalytic rate (kcat = 0.09 s-1) for trichodiene Synthase-catalyzed conversion of farnesyl diphosphate (FPP) to trichodiene at 15 °C. In a multiturnover experiment, a burst phase (kb = 4.2 s-1) corresponding to the accumulation of trichodiene on the surface of the enzyme was followed by a slower, steady-state release of products (klin = 0.086 s-1) which corresponds to kcat. These results strongly suggest that the release of trichodiene from the enzyme active site is the rate-limiting step in the overall Reaction, while the consumption of FPP is the step which limits chemical catalysis at the active site. Single-turnover experiments with trichodiene Synthase mutant D101E, for which the steady-state rate constant kcat is 1/3 of that of wild type, revealed that the mutation actually depresses t...
Kyozo Ogura - One of the best experts on this subject based on the ideXlab platform.
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Prenyltransferase Reaction involving enantiomeric discrimination. Enzymatic synthesis of ()-10, 11-epoxyfarnesol from racemic 6, 7-epoxygeranyl diphosphate and isopentenyl dipnosphate
Tetrahedron Letters, 2001Co-Authors: Hitoshi Inoue, Shinobu Ohnuma, Kyozo OguraAbstract:Abstract ( S )-10, 11-Epoxyfarnesol was formed in 66% ee from racemic 6, 7-epoxy geranyl diphosphate and isopentenyl diphosphate by farnesyl diphosphate Synthase Reaction followed by phosphatase hydrolysis.
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Farnesyl diphosphate Synthase and solanesyl diphosphate Synthase Reactions of diphosphate-modified allylic analogs: the significance of the diphosphate linkage involved in the allylic substrates for prenyltransferase.
Journal of biochemistry, 1992Co-Authors: Takeshi Gotoh, Kyozo OguraAbstract:Diphosphate-modified substrates for prenyltransferase were synthesized and examined as substrates for the prenyltransferase Reaction. They were dimethylallyl methylenediphosphonate, geranyl methylenediphosphonate, geranyl imidodiphosphate, geranyl phosphosulfate, farnesyl methylenediphosphonate, farnesyl imidodiphosphate, and farnesyl phosphosulfate. All of them except dimethylallyl methylenediphosphonate were accepted as substrates by solanesyl diphosphate Synthase to give solanesyl diphosphate and the former four analogs were also accepted as substrates by farnesyl diphosphate Synthase to give farnesyl diphosphate. The Km values of both enzymes for the methylenediphosphonate and imidodiphosphate analogs were comparable to those of the corresponding diphosphate substrates, but the phosphosulfate analogs showed much greater Km values than the diphosphate substrates. On the other hand, the Vmax values for these artificial substrates were all smaller than those for the corresponding natural substrates. Kinetic experiments with the analogs showed that the ionization-condensation-elimination mechanism proposed for the farnesyl diphosphate Synthase Reaction holds also for the solanesyl diphosphate Synthase Reaction and that the diphosphoryl structure, capable of chelating with divalent cations, is important topologically and kinetically rather than thermodynamically.
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Chain length distribution of the products formed in solanesyl diphosphate Synthase Reaction.
Journal of biochemistry, 1992Co-Authors: Shin-ichi Ohnuma, Tanetoshi Koyama, Kyozo OguraAbstract:Factors that affect the termination of isoprenoid chain elongation catalyzed by prenyltransferase were investigated. The chain-length distribution of Reaction products of solanesyl diphosphate Synthase [EC 2.5.1.11] homogeneously purified from Micrococcus luteus changed dramatically according to the concentration of the complex formed between isopentenyl diphosphate and Mg2+ (IPP-Mg) in the Reaction mixture. However, the concentration of the complex between farnesyl diphosphate and Mg2+ (FPP-Mg), the priming substrate for this Synthase, did not affect the product distribution, provided that the concentration of IPP-Mg was maintained at a certain level. Thus, the level of IPP-Mg is decisive in affecting the chain length distribution of the products of the prenyltransferase Reaction, and the Mg(2+)-dependent variability of product specificity so far observed can now be understood in terms of the effect of IPP-Mg concentration.
Karen S Anderson - One of the best experts on this subject based on the ideXlab platform.
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Pre-steady-state kinetic analysis of the trichodiene Synthase Reaction pathway.
Biochemistry, 1997Co-Authors: David E. Cane, Hsien Tai Chiu, Po-huang Liang, Karen S AndersonAbstract:The pre-steady-state kinetics of the trichodiene Synthase Reaction were investigated by rapid chemical quench methods. The single-turnover rate was found to be 3.5−3.8 s-1, a rate 40 times faster than the steady-state catalytic rate (kcat = 0.09 s-1) for trichodiene Synthase-catalyzed conversion of farnesyl diphosphate (FPP) to trichodiene at 15 °C. In a multiturnover experiment, a burst phase (kb = 4.2 s-1) corresponding to the accumulation of trichodiene on the surface of the enzyme was followed by a slower, steady-state release of products (klin = 0.086 s-1) which corresponds to kcat. These results strongly suggest that the release of trichodiene from the enzyme active site is the rate-limiting step in the overall Reaction, while the consumption of FPP is the step which limits chemical catalysis at the active site. Single-turnover experiments with trichodiene Synthase mutant D101E, for which the steady-state rate constant kcat is 1/3 of that of wild type, revealed that the mutation actually depresses t...
