Deprotonation Of C2 Research Articles

Trinuclear PdII Complexes with a C3 -Symmetric Triethynylbenzene-Based Tris-NHC Ligand: Catalytic Benefits.

A new C3 -symmetric tris-imidazolium tribromide salt 3, featuring 1,3,5-substituted triethynylbenzene, was used for the preparation of a trinuclear PdII pyridine-enhanced precatalyst preparation stabilization and initiation-type (PEPPSI) complex by triple C2 deprotonation followed by the addition of PdCl2 . Trinuclear PdII complex possessing a combination of NHC and PPh3 ligands has also been synthesized. The corresponding mononuclear palladium(II) complexes have also been synthesized for the comparison purpose. All these complexes have been characterized by using NMR spectroscopy and ESI mass spectrometry. The molecular structure of the trinuclear palladium(II) complex bearing mixed carbene and pyridine donor ligands has been established by using single crystal XRD. All the palladium(II) complexes have been used as pre-catalysts, which gave good to excellent yields in intermolecular α-arylation of 1-methyl-2-oxindole and Sonogashira coupling reaction. Catalytic studies indicate an enhanced activity of the trinuclear PdII complex in comparison to the corresponding mononuclear PdII complex for both catalytic transformations. The better performance of the trinuclear complex has also been further supported by preliminary electrochemical measurements. A negative mercury poison test was observed for both the aforementioned catalyses and therefore, it is likely that these organic transformations proceed homogeneously.

New Role of Water in Transketolase Catalysis.

Transketolase catalyzes the interconversion of keto and aldo sugars. Its coenzyme is thiamine diphosphate. The binding of keto sugar with thiamine diphosphate is possible only after C2 deprotonation of its thiazole ring. It is believed that deprotonation occurs due to the direct transfer of a proton to the amino group of its aminopyrimidine ring. Using mass spectrometry, it is shown that a water molecule is directly involved in the deprotonation process. After the binding of thiamine diphosphate with transketolase and its subsequent cleavage, a thiamine diphosphate molecule is formed with a mass increased by one oxygen molecule. After fragmentation, a thiamine diphosphate molecule is formed with a mass reduced by one and two hydrogen atoms, that is, HO and H2O are split off. Based on these data, it is assumed that after the formation of holotransketolase, water is covalently bound to thiamine diphosphate, and carbanion is formed as a result of its elimination. This may be a common mechanism for other thiamine enzymes. The participation of a water molecule in the catalysis of the one-substrate transketolase reaction and a possible reason for the effect of the acceptor substrate on the affinity of the donor substrate for active sites are also shown.

Intrinsic Strand-Incision Activity of Human UNG: Implications for Nick Generation in Immunoglobulin Gene Diversification.

Uracil arises in cellular DNA by cytosine (C) deamination and erroneous replicative incorporation of deoxyuridine monophosphate opposite adenine. The former generates C → thymine transition mutations if uracil is not removed by uracil-DNA glycosylase (UDG) and replaced by C by the base excision repair (BER) pathway. The primary human UDG is hUNG. During immunoglobulin gene diversification in activated B cells, targeted cytosine deamination by activation-induced cytidine deaminase followed by uracil excision by hUNG is important for class switch recombination (CSR) and somatic hypermutation by providing the substrate for DNA double-strand breaks and mutagenesis, respectively. However, considerable uncertainty remains regarding the mechanisms leading to DNA incision following uracil excision: based on the general BER scheme, apurinic/apyrimidinic (AP) endonuclease (APE1 and/or APE2) is believed to generate the strand break by incising the AP site generated by hUNG. We report here that hUNG may incise the DNA backbone subsequent to uracil excision resulting in a 3´-α,β-unsaturated aldehyde designated uracil-DNA incision product (UIP), and a 5´-phosphate. The formation of UIP accords with an elimination (E2) reaction where deprotonation of C2´ occurs via the formation of a C1´ enolate intermediate. UIP is removed from the 3´-end by hAPE1. This shows that the first two steps in uracil BER can be performed by hUNG, which might explain the significant residual CSR activity in cells deficient in APE1 and APE2.

Insights into the Progression of Labile Hb A1c to Stable Hb A1c via a Mechanistic Assessment of 2,3-Bisphosphoglycerate Facilitation of the Slow Nonenzymatic Glycation Process

Nonenzymatic glycation (NEG) of human hemoglobin (Hb A) consists of initial non covalent, reversible steps involving glucose and amino acid residues, which may also involve effector reagent(s) in the formation of labile Hb A1c (the conjugate acid of the Schiff base). Labile Hb A1c can then undergo slow, largely irreversible, formation of stable Hb A1c (the Amadori product). Stable Hb A1c is measured to assess diabetic progression after labile Hb A1c removal. This study aimed to increase the understanding of the distinctions between labile and stable Hb A1c from a mechanistic perspective in the presence of 2,3-bisphosphoglycerate (2,3-BPG). 2,3-Bisphosphoglycerate is an effector reagent that reversibly binds in the Hb A1c pocket and modestly enhances overall NEG rate. The deprotonation of C2 on labile Hb A1c in the formation of the Amadori product was previously proposed to be rate-limiting. Computational chemistry was used here to identify the mechanism(s) by which 2,3-BPG facilitates the deprotonation of C2 on labile Hb A1c. 2,3-Bisphosphoglycerate is capable of abstracting protons on C2 and the α-nitrogen of labile Hb A1c and can also deprotonate water and/or amino acid residues, therefore preparing these secondary reagents to deprotonate labile Hb A1c. Parallel reactions not leading to an Amadori product were found that include formation of the neutral Schiff base, dissociation of glucose from the protein, and cyclic glycosylamine formation. These heretofore under appreciated parallel reactions may help explain both the selective removal of labile from stable Hb A1c and the slow rate of NEG.

Excision of uracil from DNA by hSMUG1 includes strand incision and processing.

Uracil arises in DNA by hydrolytic deamination of cytosine (C) and by erroneous incorporation of deoxyuridine monophosphate opposite adenine, where the former event is devastating by generation of C → thymine transitions. The base excision repair (BER) pathway replaces uracil by the correct base. In human cells two uracil-DNA glycosylases (UDGs) initiate BER by excising uracil from DNA; one is hSMUG1 (human single-strand-selective mono-functional UDG). We report that repair initiation by hSMUG1 involves strand incision at the uracil site resulting in a 3′-α,β-unsaturated aldehyde designated uracil-DNA incision product (UIP), and a 5′-phosphate. UIP is removed from the 3′-end by human apurinic/apyrimidinic (AP) endonuclease 1 preparing for single-nucleotide insertion. hSMUG1 also incises DNA or processes UIP to a 3′-phosphate designated uracil-DNA processing product (UPP). UIP and UPP were indirectly identified and quantified by polyacrylamide gel electrophoresis and chemically characterised by matrix-assisted laser desorption/ionisation time-of-flight mass-spectrometric analysis of DNA from enzyme reactions using 18O- or 16O-water. The formation of UIP accords with an elimination (E2) reaction where deprotonation of C2′ occurs via the formation of a C1′ enolate intermediate. A three-phase kinetic model explains rapid uracil excision in phase 1, slow unspecific enzyme adsorption/desorption to DNA in phase 2 and enzyme-dependent AP site incision in phase 3.

A QM/MM study of the catalytic mechanism of SAM methyltransferase RlmN from Escherichia coli.

RlmN is a radical S-adenosylmethionine (SAM) enzyme that catalyzes the C2 methylation of adenosine 2503 (A2503) in 23S rRNA and adenosine 37 (A37) in several Escherichia coli transfer RNAs (tRNA). The catalytic reaction of RlmN is distinctly different from that of typical SAM-dependent methyltransferases that employs an SN 2 mechanism, but follows a ping-pong mechanism which involves the intermediate methylation of a conserved cysteine residue. Recently, the x-ray structure of a key intermediate in the RlmN reaction has been reported, allowing us to perform combined quantum mechanics and molecular mechanics (QM/MM) calculations to delineate the reaction details of RlmN at atomic level. Starting from the Cross-Linked RlmN C118A-tRNA complex, the possible mechanisms for both the formation and the resolution of the cross-linked species (IM2) have been illuminated. On the basis of our calculations, IM2 is formed by the attack of the C355-based methylene radical on the sp2 -hybridized C2 of the adenosine ring, corresponding to energy barrier of 14.4 kcal/mol, and the resolution of IM2 is confirmed to follow a radical fragmentation mechanism. The cleavage of C'-S' bond of mC355-A37 cross-link is in concert with the deprotonation of C2 by C118 residue, which is the rate-limiting step with an energy barrier of 17.4 kcal/mol. Moreover, the cleavage of C'-S' bond of IM2 can occur independently, that is, it does not require the loss of an electron of IM2 and the formation of disulfide bond between C355 and C118 as precondition. These findings would deepen the understanding of the catalysis of RlmN.

Chemical Mechanism of the Branched-Chain Aminotransferase IlvE from Mycobacterium tuberculosis.

The biosynthetic pathway of the branched-chain amino acids is essential for Mycobacterium tuberculosis growth and survival. We report here the kinetic and chemical mechanism of the pyridoxal 5'-phosphate (PLP)-dependent branched-chain aminotransferase, IlvE, from M. tuberculosis (MtIlvE). This enzyme is responsible for the final step of the synthesis of the branched-chain amino acids isoleucine, leucine, and valine. As seen in other aminotransferases, MtIlvE displays a ping-pong kinetic mechanism. pK values were identified from the pH dependence on V as well as V/K, indicating that the phosphate ester of the PLP cofactor, and the α-amino group from l-glutamate and the active site Lys204, play roles in acid-base catalysis and binding, respectively. An intrinsic primary kinetic isotope effect was identified for the α-C-H bond cleavage of l-glutamate. Large solvent kinetic isotope effect values for the ping and pong half-reactions were also identified. The absence of a quininoid intermediate in combination with the Dkobs in our multiple kinetic isotope effects under single-turnover conditions suggests a concerted type of mechanism. The deprotonation of C2 of l-glutamate and the protonation of C4' of the PLP cofactor happen synchronously in the ping half-reaction. A chemical mechanism is proposed on the basis of the results obtained here.

The Pseudouridine Synthases Proceed through a Glycal Intermediate.

The pseudouridine synthases isomerize (U) in RNA to pseudouridine (Ψ), and the mechanism that they follow has long been a question of interest. The recent elucidation of a product of the mechanistic probe 5-fluorouridine that had been epimerized to the arabino isomer suggested that the Ψ synthases might operate through a glycal intermediate formed by deprotonation of C2′. When that position in substrate U is deuterated, a primary kinetic isotope effect is observed, which indisputably indicates that the proposed deprotonation occurs during the isomerization of U to Ψ and establishes the mechanism followed by the Ψ synthases.

A Stereoselective Synthesis of a 3,4,5-Substituted Piperidine of Interest as a Selective Muscarinic (M1) Receptor Agonist

Syntheses of (1RS,2SR,6SR)-2-alkoxymethyl-, 2-hetaryl-, and 2-(hetarylmethyl)-7-arylmethyl-4,7-diaza-9-oxabicyclo[4.3.0]nonan-8-ones, of interest as potential muscarinic M1 receptor agonists, are described. A key step in the synthesis of (1RS,2SR,6SR)-7-benzyl-6-cyclobutyl-2-methoxymethyl-4,7-diaza-9-oxabicyclo[4.3.0]nonan-8-one, was the addition of isopropenylmagnesium bromide to 2-benzyloxycarbonylamino-3-tert-butyldimethylsilyloxy-2-cyclobutylpropanal. This gave the 4-tert-butyldimethylsilyloxymethyl-4-cyclobutyl-5-isopropenyloxazolidinone with the 5-isopropenyl and 4-tert-butyldimethylsilyloxymethyl groups cis-disposed about the five-membered ring by chelation controlled addition and in situ cyclisation. This reaction was useful for a range of organometallic reagents. The hydroboration–oxidation of (4SR,5RS)-3-benzyl-4-(tert-butyldimethylsilyloxymethyl)-4-cyclobutyl-5-(1-methoxyprop-2-en-2-yl)-1,3-oxazolidin-2-one gave (4SR,5RS)-3-benzyl-4-(tert-butyldimethylsilyloxymethyl)-4-cyclobutyl-5-[(SR)-1-hydroxy-3-methoxyprop-2-yl]-1,3-oxazolidin-2-one stereoselectively. 4,7-Diaza-9-oxabicyclo[4.3.0]nonan-8-ones with substituents at C2 that could facilitate C2 deprotonation were unstable with respect to oxazolidinone ring-opening and this restricted both the synthetic approach and choice of 2-heteroaryl substituent. The bicyclic system with a 2-furyl substituent at C2 was therefore identified as an important target. The addition of 1-lithio-1-(2-furyl)ethene to 2-benzyloxycarbonylamino-3-tert-butyldimethylsilyloxy-2-cyclobutylpropanal gave (4SR,5RS)-4-tert-butyldimethylsilyloxymethyl-4-cyclobutyl-5-[1-(2-furyl)ethenyl]-1,3-oxazolidinone after chelation controlled addition and in situ cyclisation. Following oxazolidinone N-benzylation, hydroboration at 35 °C, since hydroboration at 0 °C was unexpectedly selective for the undesired isomer, followed by oxidation gave a mixture of side-chain epimeric alcohols that were separated after SEM-protection and selective desilylation. Conversion of the neopentylic alcohols into the corresponding primary amines by reductive amination, was followed by N-nosylation, removal of the SEM-groups and cyclisation using a Mitsunobu reaction. Denosylation then gave the 2-furyloxazolidinonyl-fused piperidines, the (1RS,2SR,6SR)-epimer showing an allosteric agonistic effect on M1 receptors. Further studies resulted in the synthesis of other 2-substituted 4,7-diaza-9-oxabicyclo[4.3.0]nonan-8-ones and an analogous tetrahydropyran

Heterobimetallic Carbene Complexes by a Single-Step Site-Selective Metalation of a Tricarbene Ligand

The unsymmetrical tris(imidazolium) salt H3-1(Br)3, featuring a 1,2,4-substitution pattern of the central phenyl ring, after triple imidazolium C2 deprotonation reacts in a one-pot reaction with Pd(OAc)2 and [M(Cp*)(Cl)2]2 (M = Rh(III), Ir(III)) to yield heterobimetallic complexes [3] (M = Rh) and [4] (M = Ir), in which the Pd(II) ion is chelated by two ortho N-heterocyclic carbene (NHC) donors while the third NHC donor coordinates to the M(III) center, which orthometalates the central phenyl ring.

A Noninnocent Cyclooctadiene (COD) in the Reaction of an “Ir(COD)(OAc)” Precursor with Imidazolium Salts

The reactions between [Ir(COD)(μ-OAc)]2 and the functionalized imidazolium salt 1-mesityl-3-(pyrid-2-yl)imidazolium bromide (MesIPy·HBr) or 1-benzyl-3-(5,7-dimethylnaphthyrid-2-yl)imidazolium bromide (BnIN·HBr) at room temperature afford the COD-activated IrIII–N-heterocyclic carbene (NHC) complexes [Ir(1-κ-4,5,6-η-C8H12)(κ2C,N-MesIPy)Br] (1) and [Ir(1-κ-4,5,6-η-C8H12)(κ2C,N-BnIN)Br] (2), respectively. In contrast, the methoxy analogue [Ir(COD)(μ-OMe)]2 on reaction with MesIPy·HBr under identical conditions affords the IrI–NHC complex [Ir(COD)(κ2C,N-MesIPy)Br]. Treatment of [Ir(COD)(κ2C,N-MesIPy)Br] with sodium acetate does not lead to COD activation. Further, use of 2,2′-bipyridine (bpy) with [Ir(COD)(μ-X)]2 (X = MeO or AcO) in the presence of [nBu4N][BF4] affords exclusively [Ir(bpy)(COD)][BF4] (3). Metal-bound acetate is shown to be an essential promoter for activation of the COD allylic C–H bond. An examination of products reveals the following transformations of the precursor components: cleavage of the imidazolium C2–H and subsequent NHC metalation, metal oxidation from IrI to IrIII, and 2e reduction of COD, effectively resulting in 1-κ-4,5,6-η-C8H12 coordination to the metal. Mechanistic investigation at the DFT/B3LYP level of theory strongly suggests that NHC metalation precedes COD allylic C–H activation. Two distinct pathways have been examined which involve initial C2–H oxidative addition to the metal followed by acetate-assisted allylic C–H activation (path A) and the reverse sequence, i.e., deprotonation of C2–H by the acetate and subsequent allylic C–H oxidative addition to the metal (path B). The result is an IrIII–NHC–hydride−κ1,η2-C8H11 complex. Subsequent intramolecular insertion of the COD double bond into the metal–hydride bond followed by isomerization gives the final product. An acetate-assisted facile COD allylic C–H bond activation, in comparison to oxidative addition of the same to Ir, makes path A the favored pathway. This work thus raises questions about the innocence of COD, especially when metal acetates are used for the synthesis of NHC complexes from the corresponding imidazolium salts.

Histone-Catalyzed Cleavage of Nucleosomal DNA Containing 2-Deoxyribonolactone

Oxidized abasic sites such as 2-deoxyribonolactone (L) are produced in DNA by a variety of oxidizing agents, including potent cytotoxic antitumor natural products. 2-Deoxyribonolactone is labile under alkaline conditions, but its half-life in free DNA at pH 7.5 is approximately 1 week. Independent generation of L at defined positions within nucleosomes reveals that the histone proteins catalyze strand scission and increase the rate between 11- and ∼43-fold. Mechanistic studies indicate that DNA-protein cross-links are not intermediates en route to strand scission and that C2 deprotonation is the rate-determining step. The use of mutant histone H4 proteins demonstrates that the lysine-rich tail that is often post-translationally modified in cells contributes to the cleavage of L but is not the sole source of the enhanced cleavage rates. Consideration of DNA repair in cells suggests that L formation in nucleosomal DNA as part of bistranded lesions by antitumor antibiotics results in de facto double strand breaks, the most deleterious form of DNA damage.

Insight into the Role of the Counteranion of an Imidazolium Salt in Organocatalysis: A Combined Experimental and Computational Study

N-Heterocyclic carbenes (NHCs) can serve as very reactive nucleophilic catalysts and exhibit strong basicity. Herein, we initiate a combined experimental and computational investigation of the NHC-catalyzed ring-closing reactions of 4-(2-formylphenoxy)but-2-enoate derivatives 1 to uncover the relationship between the counteranion of an azolium salt, the nucleophilicity and basicity of the carbene species, and the catalytic performance of the carbene species by taking imidazolium salts IPr⋅HX (X=counteranion, IPr=1,3-bis(2,6-diisopropylphenyl)imidazol-2-ylidene) as the representative precatalysts. The plausible mechanisms of IPr-mediated ring-closing reactions have been investigated by using DFT calculations. The hydrogen-accepting ability, assigned as the basicity of the counteranion of IPr⋅HX and evaluated by DFT calculations, is correlated with the rate of deprotonation of C2 in IPr⋅HX, which could be monitored by the capture of the free carbene formed in situ with elemental sulfur. The deprotonation of C2 in IPr⋅HX with a more basic anion gives rise to a higher concentration of the free carbene and vice versa. At a relatively low concentration, IPr prefers to show a nucleophilic character to induce the intramolecular Stetter reaction. At a relatively high concentration, IPr primarily acts as a base to afford benzofuran derivatives. These data comprehensively disclose, for the first time, that the counteranions of azolium salts significantly influence not only the catalytic activity, but also possibly the reaction mechanism.

Emerging strategies in catalysis

In various cases the ILs provide a favourable environ-ment for catalyst stabilization and, in some cases, improvement inthe catalytic performance (activity and selectivity) as compared tothe reactions performed in classical organic solvents or water.However, C2 deprotonation of 1,3-dialkylimidazolium ILs iseasily effected by relatively weak bases

1,1′,3,3′‐Tetraalkyl‐2,2′‐biperimidinylidenes: Unexpected substituent effects on the reactivity of carbon–carbon double bond

AbstractC2 deprotonation of 1,3‐dibutylperimidinium bromide (1a) with sodium hydride and a catalytic amount of potassium tert‐butoxide in dry THF led to the formation of the exceptionally inert tetraaminoalkene 2a. In contrast, isostructural tetrakis(2‐methoxyethyl)‐tetraaminoalkene (2b) instantaneously reacted with O2 to yield urea 3b, and silver nitrate was readily reduced with 2b to form a silver mirror. Compound 2a has been characterized by X‐ray diffraction studies; the naphtho‐pyrimidine skeleton imposes structural constraints and some rigidity to the CC bonding. © 2003 Wiley Periodicals, Inc. Heteroatom Chem 14:82–87, 2003; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/hc.10088

Activation of thiamine diphosphate in pyruvate decarboxylase from Zymomonas mobilis

Replacement of tryptophan 392 located in the active site cavity of pyruvate decarboxylase (PDC; EC 4.1.1.1) from Zymomonas mobilis by methionine or glutamine yields enzymes with smaller catalytic constants of 8.5 s −1 and 3.6 s −1 at 4°C, compared to that of the wild-type enzyme (17 s −1). The rate constants of the H/D exchange at the C2 of the coenzyme thiamine diphosphate have been determined to be 130 s −1 for the wild-type enzyme, 56 s −1 for the methionine and 30 s −1 for the glutamine mutant, respectively. A group with a p K a of about 5 has been identified to be essential for C2 deprotonation of the enzyme-bound thiamine diphosphate from the pH dependence of the H/D exchange.