Enolimine Tautomer Research Articles

Exploring the factors influencing the ketoenamine-enolimine tautomeric equilibrium of pyridoxal 5′-phosphate in branched-chain aminotransferases

Pyridoxal 5′-phosphate (PLP), the active form of vitamin B6, is a critical coenzyme for various enzymes. It generates a Schiff base with the substrate and exhibits ketoenamine and enolimine tautomeric forms due to intramolecular proton transfer. This study aims to ascertain the predominant tautomeric form of the PLP Schiff base in MtIlvE and analyze its influencing factors. Molecular dynamics simulations indicate that the ketoenamine tautomer has higher binding free energy than the enolimine tautomer. Density functional theory calculations suggest that, despite their ability to interconvert at a relatively low energy barrier, the ketoenamine tautomer is thermodynamically more stable. Factors affecting the keto-enol tautomeric equilibrium were investigated by constructing various QM-cluster models. Our results demonstrate that both the protonation of the pyridine nitrogen and the presence of Tyr209, which stabilizes the O3 anion, shift the tautomeric equilibrium toward the ketoenamine configuration. These findings provide a theoretical basis for investigating enzyme catalytic mechanisms.

Solvent-free one-pot two component synthesis. Tuning of optoelectronic properties of fluorescent phloroglucinol-aryl Schiff base compounds

A series of new aryl-Schiff base derivatives were synthetized from the one-pot approach: two component condensation reaction in absence of solvent, in quantitative yields with a high atom economy under green conditions. All of the derivatives were fully characterized by NMR, vibrational absorption analysis, UV/vis absorption and fluorescence spectroscopy, and high-resolution mass spectrometry. The aryl-Schiff bases predominantly present the enol-imine tautomer in acetone-d6, while both enol-imine and keto-imine tautomer readily concur in dimethyl sulphoxide-d6. The emission of the series could be tailored by substituting electron donor or electron acceptor groups in the aryl-Schiff bases from fluorescent to not fluorescent molecules, respectively. Fluorescent molecules emit blue, cyan and yellow color with quantum yields in the 2.4–6.2 % range and average lifetime of ∼0.2 ns. The large Stokes’ shift for the molecule bearing a methylalcoxy (3248 cm−1) and diethyl amine (5017 cm−1) suggests a possible intramolecular charge transfer process as also corroborated by cyclic voltammetry, and explaining their low fluorescence quantum yield. The aryl-Schiff base substituted with the methylalcoxy (2b) was used as an electron donor material in photovoltaic devices in configuration: ITO/PEDOT:PSS/2b:PC61BM/Fields metal. The device built under atmospheric conditions displayed an intrinsic power conversion efficiency of PCE ∼0.4 %, with a Voc of 420 mV and Jsc 4.28 mA/cm2.

Novel Schiff Bases of C-Methylresorcinarene Derivatives

The article presents the synthesis and properties of two new Schiff bases of resorcinarene derivatives. The Schiff bases were obtained by the reaction of formylresorcinarene with aromatic (o-aminophenol) and aliphatic (N,N-dimethyldiaminoethane) amines in chloroform. The synthesized Schiff bases exist in equilibrium of several tautomers, as evident from the IR, UV, NMR spectra and cyclic voltammetry data analysis. In DMF, methanol, and acetonitrile, the tautomeric equilibrium is shifted toward the enol-imine tautomers.

Push-pulling induces the excited-state intramolecular proton transfer in 2′-aminochalcones

Usually, chalcones are not luminescent, although their derivatives with push-pull patterns of substitution may present fair quantum yields. Intramolecular hydrogen bonding between a hydrogen donor (–OH or –NH) and the chalcone carbonyl promotes Excited State Intramolecular Proton Transfer (ESIPT). If the dye is fluorescent, the emission can derive from one of the excited tautomers, or from both and in this case a dual emission is observed. Here, 2′-aminochalcones with a strong push-pull character, incorporating electron-donating groups on one side, and electron-withdrawing groups on the amine, present dual emission ascribed to ESIPT between the keto-enamine and the enol-imine tautomers. The hydrogen bonds strength was studied by NMR and DFT calculations. A correlation between the tautomerization energy gap and the hydrogen bond strength was observed, depending on the electron donating or electron withdrawing group on the amine. The electron withdrawing groups bonded to the proton donor have little influence on the quantum yield and emission wavelength. In contrast, the electron donating groups connected to the proton acceptor have a significant effect on the quantum yield and induce an absorption and emission red shift.

Naphthalene-based bis-N-salicylidene aniline dyes: Crystal structures, Hirshfeld surface analysis, computational study and molecular docking with the SARS-CoV-2 proteins

In this work, we report structural and computational studies of a series of naphthalene-based bis-N-salicylidene aniline dyes, namely N,N′-bis-salicylidene-1,5-diaminonaphthalene (1), N,N′-bis(3-hydroxysalicylidene)-1,5-diaminonaphthalene (2) and N,N′-bis(3-methoxysalicylidene)-1,5-diaminonaphthalene (3). For 3, two polymorphs are known, namely 3red and 3yellow. Both polymorphs of 3 were analyzed and discussed. All the molecules adopt an enol-imine tautomer, stabilized by two intramolecular O–HcdotsN hydrogen bonds. The structure of 2 is further stabilized by a couple of additional O–HcdotsO hydrogen bonds and by intermolecular O–HcdotsO interactions, yielding a 1D zig-zag supramolecular chain. Molecules of 2, 3red and 3yellow are interlinked through intermolecular C–Hcdotsπ interactions, while the crystal packing of 1 and 2 is also described by intermolecular πcdotsπ interactions. More than 90% of the total Hirshfeld surface area for all the discussed molecules is occupied by HcdotsH, HcdotsC, HcdotsO and CcdotsC contacts. The polymorphs 3red and 3yellow, despite being chemically the same, differ geometrically, thus yielding remarkably different Hirshfeld surfaces. The Hirshfeld surface of 3yellow is very similar to that of 2. All structures are mainly characterized by the dispersion energy framework followed by the less significant electrostatic energy framework contribution. Molecular docking studies were employed to inspect the effect of 1–3 on the SARS-CoV-2 protein targets. The docking analysis revealed that the dye 2 showed the best binding energies toward Papain-like protease (PLpro, –10.40 kcal/mol), nonstructural protein 14 (nsp14 (N7-MTase), –10.10 kcal/mol), RdRp-RTP (–9.70 kcal/mol) and nonstructural protein 3 (nsp3_range 207-379-MES, –9.30 kcal/mol). The obtained results can give an insight into chemical and biological activities of the studied molecules that could aid in designing of potent reagents SARS-CoV-2.Supplementary InformationThe online version contains supplementary material available at 10.1007/s13738-021-02438-y.

Theoretical evidence for the resonance-inhibited hydrogen bonding (RIHB) in enol-imine tautomers

• Resonance inhibited hydrogen bonds (RIHBs) can exist in some of enol-imine tautomers. • Generally in an RIHB system, π-resonance attempts to decrease the hydrogen bond strength, while σ-planarity can more or less compensate for this impairment. • Π-electron donating groups (EDGs) can strengthen an RIHB by hindering the resonance flow in a part of the molecule. • The ability of an EDG to tune an RIHB is enhanced with increasing the magnitude of its Hammet resonance constant. The theory of resonance-inhibited hydrogen bonding (RIHB) is tested on some illustrative examples with enol-imine tautomerism. In contrast to the resonance-assisted hydrogen bonding (RAHB), theoretical descriptors show that here, π-resonance weakens the intramolecular hydrogen bond (IMHB) strength in the given structures. The results show that it is possible to tune the strength of RIHB by substitution in a suitable position in the molecule. Once a substituent hiders the original resonance in the RIHB system, it leads to reinforcement of IMHB and vice versa. These effects mostly are enhanced with increasing the magnitude of Hammet resonance constants. Interestingly, the IMHB becomes 0.08 Å shorter, and thus quite stronger by using a powerful π-electron donating group like –O − on the basis of the RIHB theory. In general, RIHB and RAHB complement each other well, and provide a very good insight into the interplay between the resonance and hydrogen bonds.

Tautomeric Equilibria in Solutions of 2-Phenacylbenzimidazoles

Detailed NMR spectral analysis of DMSO-d6 solutions of the series of substituted 2-phenacylbenzimidazoles (ketimine form, K) reveals two from three tautomeric forms. Integrals of the 1H NMR signals are used in establishing the molar ratio of tautomers. The experimental analyses are supported by quantum-chemical calculations, which satisfactorily reproduced the experimental trends. Although the reported semiempirical quantum-chemical calculations show that enaminone E, i.e., 2-(1,3-dihydro-2H-benzo[d]imidazol-2-ylidene)-1-phenylethan-1-one, was thermodynamically most stable, the results of MP2 ab initio calculations reveal the following order of stability: ketimine > enolimine > enaminone (substituents do not affect this sequence). 13C CPMAS NMR spectral data reveal that in the crystalline state the enolimine tautomer O is predominant in the p-CH3 and p-NO2 substituted congeners.

The Rich Tautomeric Behavior of Campestarenes.

Campestarenes are a new family of Schiff-base macrocycles that form selectively in a one-step synthesis. These macrocycles with five-fold symmetry show solvent-dependent tautomerization and dimerization or aggregation. In this paper, we have prepared new soluble campestarenes that do not aggregate. The initial single-crystal X-ray diffraction study of a campestarene reveals that these macrocycles are nearly flat. The tautomeric behavior of the campestarenes has been extensively studied by variable-temperature, multinuclear NMR spectroscopy, UV/Vis spectroscopy, and IR spectroscopy. In polar solvents, such as DMF, the molecules exist predominantly in their keto-enamine form, but the enol-imine tautomer is dominant in non-polar solvents. A detailed computational study of the tautomeric forms of campestarenes provides a theoretical basis for their behavior and corroborates the experimental data. The results of this study give the first comprehensive understanding of the electronic and spectroscopic properties of these pentagonal macrocycles.

Two fluorescent Schiff base sensors for Zn(2+): the Zn(2+)/Cu(2+) ion interference.

Two simple and low cost 2,4-di-tert-butyl-6-[(1-hydroxycyclohexylmethylimino)methyl]phenol (L1) and 2-[{(1-hydroxycyclohexyl)methylimino}methyl]phenol (L2) Schiff base sensors exhibiting selectivity for Zn(2+) in water:methanol (95:5, v/v, 10 mM HEPES) are described. L1 and L2 display an "off-on" fluorescence effect forming the L1·Zn and L2·Zn complexes, respectively. In the case of L1·Zn, the emission response is quenched by the addition of Cu(2+) forming the respective L1·Cu complex; in spite of that, the fluorescence signal can be completely restored only by the addition of tartrate anions (C4H4O6(2-)) forming again L1·Znvia the "off-on" displacement approach. However, in the case of L2·Zn no Cu(2+) interference is observed, which is a typical problem for Zn(2+) sensors. Here we describe that a very subtle structural change in the ligand during transition from the enol-imine tautomer in L1 to the keto-enamine tautomer in L2 is enough to modulate the Zn(2+)/Cu(2+) selectivity. Also, the Zn(2+)vs. Cd(2+) discrimination for L1 and L2 is proved. Moreover, we found that the interaction between both L·Zn complexes and tartrate anions completely restored the free ligands by the ligand substitution mechanism even in a more efficient association than phosphate anions. Further, a second colorimetric response channel upon addition of Fe(2+) was observed for L1 and L2. Then, TD-DFT theoretical calculations were conducted in order to study the efficiency of the sensors to give different responses in the presence of such metal ions. Finally, the L2 sensor successfully detects Zn(2+) in Jurkat cells cultured with and without Zn(2+) enriched medium.

Lyotropic liquid crystallinity in mixed-tautomer Schiff-base macrocycles.

Schiff-base macrocycles that combine keto-enamine and enol-imine tautomers within the same ring were synthesized and characterized. These new macrocycles form host-guest complexes with organic cations and unexpectedly form a lyotropic liquid crystal in organic solvents such as chloroform and toluene.

A hybrid QM/MM simulation study of intramolecular proton transfer in the pyridoxal 5'-phosphate in the active site of transaminase: influence of active site interaction on proton transfer.

Pyridoxal 5'-phosphate (PLP) Schiff base, a versatile cofactor, exhibits a tautomeric equilibrium that involves an intramolecular proton transfer between the N-protonated zwitterionic ketoenamine tautomer and the O-protonated covalent enolimine tautomer. It has been postulated that for the catalytic activity, the PLP has to be in the zwitterionic ketoenamine tautomeric form. However, the exact position of the tautomeric equilibrium of Schiff base in the active site of PLP-dependent enzyme is not known yet. In the present work, we investigated the tautomeric equilibrium for the external aldimine state of PLP aspartate (PLP-Asp) Schiff base in the active site of aspartate aminotransferase (AspAT) using combined quantum mechanical and molecular mechanical simulations. The main focus of the present study is to analyze the factors that control the tautomeric equilibrium in the active sites of various PLP-dependent enzymes. The results show that the ketoenamine tautomer is more preferred than the enolimine tautomer both in the gas and aqueous phases as well as in the active site of AspAT. Current simulations show that the active site of AspAT is more suitable for the ketoenamine tautomer compared to the enolimine tautomer. Interestingly, the Tyr225 acts as a proton donor to the phenolic oxygen in the ketoenamine tautomer, while in the covalent enolimine tautomer, it acts as a proton acceptor to the phenolic oxygen. Finally, the metadynamics study confirms this result. The calculated free energy barrier is about 7.5 kcal/mol. A comparative analysis of the microenvironment created by the active site residues of three different PLP-dependent enzymes (aspartate aminotransferase, Dopa decarboxylase, and Ala-racemase) has been carried out to understand the controlling factor(s) of the tautomeric equilibrium. The analysis shows that the intermolecular hydrogen bonding between active site residues and the phenolic oxygen of PLP shifts the tautomeric equilibrium toward the N-protonated ketoenamine tautomeric form.

NMR Studies of Protonation and Hydrogen Bond States of Internal Aldimines of Pyridoxal 5′-Phosphate Acid–Base in Alanine Racemase, Aspartate Aminotransferase, and Poly-l-lysine

Using (15)N solid-state NMR, we have studied protonation and H-bonded states of the cofactor pyridoxal 5'-phosphate (PLP) linked as an internal aldimine in alanine racemase (AlaR), aspartate aminotransferase (AspAT), and poly-L-lysine. Protonation of the pyridine nitrogen of PLP and the coupled proton transfer from the phenolic oxygen (enolimine form) to the aldimine nitrogen (ketoenamine form) is often considered to be a prerequisite to the initial step (transimination) of the enzyme-catalyzed reaction. Indeed, using (15)N NMR and H-bond correlations in AspAT, we observe a strong aspartate-pyridine nitrogen H-bond with H located on nitrogen. After hydration, this hydrogen bond is maintained. By contrast, in the case of solid lyophilized AlaR, we find that the pyridine nitrogen is neither protonated nor hydrogen bonded to the proximal arginine side chain. However, hydration establishes a weak hydrogen bond to pyridine. To clarify how AlaR is activated, we performed (13)C and (15)N solid-state NMR experiments on isotopically labeled PLP aldimines formed by lyophilization with poly-L-lysine. In the dry solid, only the enolimine tautomer is observed. However, a fast reversible proton transfer involving the ketoenamine tautomer is observed after treatment with either gaseous water or gaseous dry HCl. Hydrolysis requires the action of both water and HCl. The formation of an external aldimine with aspartic acid at pH 9 also produces the ketoenamine form stabilized by interaction with a second aspartic acid, probably via a H-bond to the phenolic oxygen. We postulate that O-protonation is an effectual mechanism for the activation of PLP, as is N-protonation, and that enzymes that are incapable of N-protonation employ this mechanism.

Tautomerism in Quinoxalines Derived from the 1,4-Naphthoquinone Nucleus: Acid Mediated Synthesis, X-ray Molecular Structure of 5-Chlorobenzo[f]quinoxalin-6-ol and Density Functional Theory Calculations

The reaction of tert-butyl 2-(3-chloro-1,4-dioxo-1,4-dihydronaphthalen-2-ylamino) ethylcarbamate with CF3COOH/CH2Cl2 yields 5-chloro-3,4-dihydrobenzo[f]quinoxalin6(2H)-one which undergoes acid-promoted dehydrogenation in the presence of water to give novel 5-chlorobenzo[f]quinoxalin-6-ol. The molecular structure of 5-chlorobenzo[f]quinoxalin6-ol in the solid state, determined by an X-ray diffraction (XRD) study, and the solution data confirm that it exists as the enol-imine tautomer, both in the solid state and in solution, differently from 5-chloro-3,4-dihydrobenzo[f]quinoxalin-6(2H)-one, which exhibits the keto-amine arrangement. Density functional theory (DFT) calculations confirmed the preference of 5-chlorobenzo[f]quinoxalin-6-ol and of the derivatives containing H and CH3 groups in place of the Cl atom for the enol-imine tautomer. It is suggested that the enol-imine structure is preferred for 5-chlorobenzo[f]quinoxalin-6-ol as a consequence of the higher aromatic character of this structure in comparison with the keto-amine form. DFT calculations carried out on the two tautomers of the benzo[a]phenazin-5(7H)-ones analogous to the benzo[f]quinoxalin-6(4H)-ones showed that the relative stabilities are dominated by solvation effects in the first case and the degree of aromaticity, in the latter.

(E)-1-[(3-Iodophenyl)iminomethyl]naphthalen-2-ol

In the title mol­ecule, C17H12INO, the dihedral angle between the naphthaldeyde plane and the 3-iodo­aniline plane is20.07 (13)°. It exists in the solid state as an enol–imine tautomer with a strong intra­molecular O—H⋯N hydrogen bond.

Synthesis, Characterization, and Tautomeric Properties of Some Azo-azomethine Compounds

The primary azo compound 1-(3-formyl-4-hydroxyphenylazo)-4-nitrobenzene reacts with some aliphatic and aromatic diamines and yields the corresponding azo-azomethine compounds. These compounds were characterized by elemental analysis, IR, UV/Vis, and NMR spectroscopy. The primary azo compound exists entirely in the azo form in solution as well as in the solid phase. The tautomeric structure of azo-azomethine compounds heavily depends on the solvent and the substituents. Aliphatic diamine-based compounds favor the enol-imine tautomer while aromatic diamine-based compounds have structures that lie between the two enol-imine and keto-amine tautomers due to a relatively strong intramolecular hydrogen bond. The compounds exhibit positive solvatochromism (bathochromic shift) so that their absorption bands move toward longer wavelengths as the polarity of the solvents increases. In addition, UV/Vis spectrophotometry has shown that the studied compounds have molar extinction coefficients larger than 40000.

Theoretical Study of the Proton Transfer in Enaminones

Problem statement: Hydrogen bonding has a vital rule to unrevealed the nature of many different interactions both in gas phase and condensed media thus it is one of the most important concepts in chemistry. Enaminoes with their ability to form intrahydrogen bonded cheated rings represent one of the suitable compounds to study such concept. Approach: The three possible tautomers of eneminones were fully optimized at several theoretical levels including B3LYP with the 6-31+G(d,p), 6-311++G(d,p) and aug-cc-pVDZ basis sets as well as MP2/6-311++G(d,p) level of theory. Then a search for the possible transition state between the two most stable tautomers; the ketamine and the enolimine was done at the B3LYP/6-311++G(d,p) level of theory. Results: The ketamine is the most stable while the diketo is the less stable within the tautomeric mixture and the proton transfer occurs through a transition state that is energetically resemble the enolimine tautomer. Conclusion: The interconversion of the tautomers is preferable in the enolimine → ketamine direction.

2-{[(4-{[(2-Hydroxyphenyl)(phenyl)methylidene]amino}phenyl)imino](phenyl)methyl}phenol

The title mol­ecule, C32H24N2O2, has a crystallographically imposed inversion centre and exists in the crystal as an enol–imine tautomer. The mol­ecular structure is stabilized by two strong intra­molecular O—H⋯N hydrogen bonds. The dihedral angles between the central benzene ring and the mean planes of the phenyl substituents are 59.99 (1) and 62.79 (2)°. In the crystal, the mol­ecules are arranged into (010) layers via C—H⋯π inter­actions.

2-[1-(3-Aminophenylimino)ethyl]phenol

The title compound, C14H14N2O, exists as the enol–imine tautomer. A strong intra­molecular hydrogen bond between O and N atoms forms a six-membered ring with an S(6) graph-set motif, which is approximately coplanar with the phenol ring, the inter­planar angle being 3.4 (3)°. In the crystal, inter­molecular C—H⋯O hydrogen bonds and N—H⋯π inter­actions link the mol­ecules into infinite chains along [100].

Identification of the Structural Determinants for the Stability of Substrate and Aminoacrylate External Schiff Bases in O-Acetylserine Sulfhydrylase-A

O-Acetylserine sulfhydrylase is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that catalyzes the final step in the cysteine biosynthetic pathway in enteric bacteria and plants, the replacement of the beta-acetoxy group of O-acetyl-L-serine (OAS) by a thiol to give L-cysteine. Previous studies of the K41A mutant enzyme showed L-methionine bound in an external Schiff base (ESB) linkage to PLP as the enzyme was isolated. The mutant enzyme exists in a closed form, optimizing the orientation of the cofactor PLP and properly positioning active site functional groups for reaction. The trigger for closing the active site upon formation of the ESB is thought to be interaction of the substrate alpha-carboxylate with the substrate-binding loop comprised of T68, S69, G70, and N71, and Q142, which is positioned above the cofactor as one looks into the active site. To probe the contribution of these residues to the active site closing and orientation of PLP in the ESB, T68, S69, N71, and Q142 were changed to alanine. Absorbance, fluorescence, near UV-visible CD, and (31)P NMR spectral studies and pre-steady state kinetic studies were used to characterize the mutant enzymes. All of the mutations affect closure of the active site, but to differing extents. In addition, the site appears to be more hydrophilic given that the ESBs do not exhibit a significant amount of the enolimine tautomer. No buildup of the alpha-aminoacrylate intermediate (AA) is observed for the T68A and Q142A mutant enzymes. However, pyruvate is produced at a rate much greater than that of the wild-type enzyme. Data suggest that T68 and Q142 play a role in stabilizing the AA. Both residues donate a hydrogen bond to one of the carboxylate oxygens of the methionine ESB and may also be responsible for the proper orientation of the ESB to generate the AA. The S69A and N71A mutants exhibit formation of the AA, but the rate constant for its formation from the ESB is decreased by 1 order of magnitude compared to that of the wild type. S69 donates a hydrogen bond to the substrate carboxylate in the ESB, while N71 donates hydrogen bonds to O3' of the cofactor and the carboxylate of the ESB; these side chains may also affect the orientation of the ESB. Data suggest that interaction of intermediates with the substrate-binding loop and Q142 gives a properly aligned Michaelis complex and facilitates the beta-elimination reaction.

(E)-3-[(4-Butyl-phen-yl)imino-meth-yl]benzene-1,2-diol.

The title compound, C17H19NO2, exists as an enol–imine tautomer. The dihedral angle between the two benzene rings is 4.6 (2)°. The mol­ecular structure is stabilized by intramol­ecular O—H⋯O and O—H⋯N hydrogen bonds which generate S(5) and S(6) ring motifs, respectively. In the crystal, mol­ecules are linked into centrosymmetric dimers by pairs of O—H⋯O hydrogen bonds. In addition, C—H⋯π inter­actions involving both benzene rings are observed.