RESUMEN
A naringinase complex was chemically aminated prior to its immobilization on glyoxyl-agarose to develop a robust biocatalyst for juice debittering. The effects of amination on the optimal pH and temperature, thermal stability, and debittering performance were analyzed. Concentration of amino groups on catalysts surface increased in 36 %. Amination reduced the ß-glucosidase activity of naringinase complex; however, did not affect optimal pH and temperature of the enzyme and it favored immobilization, obtaining α-l-rhamnosidase and ß-d-glucosidase activities of 1.7 and 4.2 times the values obtained when the unmodified enzymes were immobilized. Amination favored the stability of the immobilized biocatalyst, retaining 100 % of both activities after 190 h at 30 °C and pH 3, while its non-aminated counterpart retained 80 and 52 % of α-rhamnosidase and ß-glucosidase activities, respectively. The immobilized catalyst showed a better performance in grapefruit juice debittering, obtaining a naringin conversion of 7 times the value obtained with the non-aminated catalyst.
Asunto(s)
Enzimas Inmovilizadas , Jugos de Frutas y Vegetales , Glioxilatos , Sefarosa , Jugos de Frutas y Vegetales/análisis , Enzimas Inmovilizadas/química , Enzimas Inmovilizadas/metabolismo , Aminación , Concentración de Iones de Hidrógeno , Sefarosa/química , Glioxilatos/química , Citrus/química , Citrus/enzimología , Estabilidad de Enzimas , Biocatálisis , Complejos Multienzimáticos/química , Complejos Multienzimáticos/metabolismo , Glicósido Hidrolasas/química , Glicósido Hidrolasas/metabolismo , beta-Glucosidasa/química , beta-Glucosidasa/metabolismo , Temperatura , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Flavanonas/química , Flavanonas/metabolismo , CatálisisRESUMEN
The influence and its mechanism of ultrasound on acetaldehyde/glyoxylic acid competing bridged the polymerization coloration of flavan-3-ols in model wine solution were investigated by high-performance liquid chromatography-mass spectrometry (HPLC-MS) and kinetics and thermodynamic model. The results indicate that ultrasound could significantly accelerate the polymerization coloration and further modify wine color. In addition, the polymerization reaction conformed first-order reaction model, and the reaction rate constant (k) values were markedly increased by ultrasound, accelerating the coloration reaction, especially in the model wine containing glyoxylic acid. Besides, the polymerization processing was non-spontaneous and endothermic according to the thermodynamic analysis. In conclusion, ultrasound was indeed conducive to accelerate glyoxylic acid/acetaldehyde-bridged the polymerization of flavan-3-ols and further effect the wine color, which could provide a theoretical basis for the scientific analysis of the mechanism of ultrasound modifying wine color.
Asunto(s)
Vino , Vino/análisis , Acetaldehído/análisis , Glioxilatos/química , Cromatografía Líquida de Alta Presión , Polifenoles/análisis , Polímeros/análisis , Flavonoides/análisisRESUMEN
We herein describe a C4 sulfonylmethylation of pyrazol-5-amines with glyoxylic acid and sodium sulfinates. The reaction only needed water as a solvent, and it featured mild reaction conditions, simple operation, and high regioselectivity. Various C4 sulfonylmethylated pyrazol-5-amines were obtained in good to excellent yields. Moreover, this sulfonylmethylation method was applicable for C(sp2)-H sulfonylmethylation of other substrates such as enamines, indoles, and antipyrines by adding a catalyst and changing the solvent. Biological evaluation revealed that some products had antiproliferative activity against cancer cell lines.
Asunto(s)
Aminas/síntesis química , Antineoplásicos/síntesis química , Glioxilatos/química , Pirazoles/síntesis química , Ácidos Sulfínicos/química , Aminas/química , Aminas/farmacología , Antineoplásicos/química , Antineoplásicos/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Metilación , Estructura Molecular , Pirazoles/química , Pirazoles/farmacologíaRESUMEN
In this study, to investigate the influence of glyoxylate bearing Schiff base on bioactivity to chitosan quaternary ammonium salts, different chitosan derivatives were synthesized by ion exchange of glyoxylate bearing Schiff base with chitosan quaternary ammonium salts (TMCI and HACC). For this purpose, glyoxylate was prepared by Schiff base reaction of glyoxylic acid and amino heterocycles and it was further ionization to substitute iodide ions and chloride ions. After structural characterization by FTIR and 1H NMR, the antifungal and antioxidant activities were measured. Results indicated that glyoxylate bearing Schiff base could improve the bioactivity of TMCI and HACC obviously. Specifically, anionic TMCI with Schiff base of amino pyridines possessed best antioxidant activity >92.40% at 1.6 mg/mL against DPPH radicals. Meanwhile, they showed antifungal activity >84.88% at 1.0 mg/mL against G. cingulate. Furthermore, the cytotoxicity was evaluated, and all samples showed good cell viability >80.14% at 1000 µg/mL.
Asunto(s)
Antifúngicos/farmacología , Antioxidantes/farmacología , Quitosano/farmacología , Hongos/efectos de los fármacos , Glioxilatos/farmacología , Compuestos de Amonio Cuaternario/farmacología , Antifúngicos/síntesis química , Antifúngicos/química , Antioxidantes/síntesis química , Antioxidantes/química , Compuestos de Bifenilo/antagonistas & inhibidores , Quitosano/química , Glioxilatos/química , Pruebas de Sensibilidad Microbiana , Picratos/antagonistas & inhibidores , Compuestos de Amonio Cuaternario/química , Bases de Schiff/química , Bases de Schiff/farmacologíaRESUMEN
The isocitrate lyase paralogs of Mycobacterium tuberculosis (ICL1 and 2) are essential for mycobacterial persistence and constitute targets for the development of antituberculosis agents. We report that (2R,3S)-2-hydroxy-3-(nitromethyl)succinic acid (5-NIC) undergoes apparent retro-aldol cleavage as catalyzed by ICL1 to produce glyoxylate and 3-nitropropionic acid (3-NP), the latter of which is a covalent-inactivating agent of ICL1. Kinetic analysis of this reaction identified that 5-NIC serves as a robust and efficient mechanism-based inactivator of ICL1 (kinact/KI = (1.3 ± 0.1) × 103 M-1 s-1) with a partition ratio <1. Using enzyme kinetics, mass spectrometry, and X-ray crystallography, we identified that the reaction of the 5-NIC-derived 3-NP with the Cys191 thiolate of ICL1 results in formation of an ICL1-thiohydroxamate adduct as predicted. One aspect of the design of 5-NIC was to lower its overall charge compared to isocitrate to assist with cell permeability. Accordingly, the absence of the third carboxylate group will simplify the synthesis of pro-drug forms of 5-NIC for characterization in cell-infection models of M. tuberculosis.
Asunto(s)
Inhibidores Enzimáticos/química , Isocitratoliasa/antagonistas & inhibidores , Mycobacterium tuberculosis/enzimología , Succinatos/química , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/metabolismo , Glioxilatos/química , Glioxilatos/metabolismo , Isocitratoliasa/química , Isocitratoliasa/metabolismo , Cinética , Modelos Químicos , Nitrocompuestos/química , Nitrocompuestos/metabolismo , Propionatos/química , Propionatos/metabolismo , Unión Proteica , Succinatos/síntesis química , Succinatos/metabolismoRESUMEN
Low-temperature plasma is being widely used in the various fields of life science, such as medicine and agriculture. Plasma-activated solutions have been proposed as potential cancer therapeutic reagents. We previously reported that plasma-activated Ringer's lactate solution exhibited selective cancer-killing effects, and that the plasma-treated L-sodium lactate in the solution was an anti-tumor factor; however, the components that are generated through the interactions between plasma and L-sodium lactate and the components responsible for the selective killing of cancer cells remain unidentified. In this study, we quantified several major chemical products, such as pyruvate, formate, and acetate, in plasma-activated L-sodium lactate solution by nuclear magnetic resonance analysis. We further identified novel chemical products, such as glyoxylate and 2,3-dimethyltartrate, in the solution by direct infusion-electrospray ionization with tandem mass spectrometry analysis. We found that 2,3-dimethyltartrate exhibited cytotoxic effects in glioblastoma cells, but not in normal astrocytes. These findings shed light on the identities of the components that are responsible for the selective cytotoxic effect of plasma-activated solutions on cancer cells, and provide useful data for the potential development of cancer treatments using plasma-activated L-sodium lactate solution.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Glioblastoma/metabolismo , Gases em Plasma/química , Lactato de Sodio/química , Tartratos/toxicidad , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Formiatos/química , Glioxilatos/química , Humanos , Ácido Pirúvico/química , Tartratos/químicaRESUMEN
Tris is an extensively used buffer that presents a primary amine group on its structure. In the present work trypsin, chymotrypsin and penicillin G acylase (PGA) were immobilized/stabilized on glyoxyl agarose in presence of different concentrations of Tris (from 0 to 20 mM). The effects of the presence of Tris during immobilization were studied analyzing the thermal stability of the obtained immobilized biocatalysts. The results indicate a reduction of the enzyme stability when immobilized in the presence of Tris. This effect can be observed in inactivations carried out at pH 5, 7, and 9 with all the enzymes assayed. The reduction of enzyme stability increased with the Tris concentration. Another interesting result is that the stability reduction was more noticeable for immobilized PGA than in the other immobilized enzymes, the biocatalysts prepared in presence of 20 mM Tris lost totally the activity at pH 7 just after 1 h of inactivation, while the reference at this time still kept around 61 % of the residual activity. These differences are most likely due to the homogeneous distribution of the Lys groups in PGA compared to trypsin and chymotrypsin (where almost 50% of Lys group are in a small percentage of the protein surface). The results suggest that Tris could be affecting the multipoint covalent immobilization in two different ways, on one hand, reducing the number of available glyoxyl groups of the support during immobilization, and on the other hand, generating some steric hindrances that difficult the formation of covalent bonds.
Asunto(s)
Enzimas Inmovilizadas/química , Glioxilatos/química , Penicilina Amidasa/química , Sefarosa/química , Trometamina/química , Tripsina/química , Tampones (Química) , Estabilidad de Enzimas , Concentración de Iones de HidrógenoRESUMEN
During the glyoxylate cycle, isocitrate lyases (ICLs) catalyze the lysis of isocitrate to glyoxylate and succinate. Itaconate has been reported to inhibit an ICL from Mycobacterium tuberculosis (tbICL). To elucidate the molecular mechanism of ICL inhibition, we determined the crystal structure of tbICL in complex with itaconate. Unexpectedly, succinate and itaconate were found to bind to the respective active sites in the dimeric form of tbICL. Our structure revealed the active site architecture as an open form, although the substrate and inhibitor were bound to the active sites. Our findings provide novel insights into the conformation of tbICL upon its binding to a substrate or inhibitor, along with molecular details of the inhibitory mechanism of itaconate.
Asunto(s)
Isocitratoliasa/química , Isocitratoliasa/metabolismo , Isocitratos/química , Succinatos/química , Succinatos/metabolismo , Ácido Succínico/química , Ácido Succínico/metabolismo , Catálisis , Dominio Catalítico/fisiología , Glioxilatos/química , Mycobacterium tuberculosis/química , Mycobacterium tuberculosis/metabolismo , Conformación ProteicaRESUMEN
The isocitrate lyases (ICL1/2) are essential enzymes of Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis. At present, no ICL1/2 inhibitors have progressed to clinical evaluation, despite extensive drug discovery efforts. Herein, we surveyed succinate analogs against ICL1 and found that dicarboxylic acids constrained in their synperiplanar conformations, such as maleic acid, comprise uncompetitive inhibitors of ICL1 and inhibit more potently than their trans-isomers. From this, we identified cis-2,3 epoxysuccinic acid (cis-EpS) as a selective, irreversible covalent inactivator of Mtb ICL1 (kinact/Kinact= (5.0 ± 1.4) × 104 M-1 s-1; Kinact = 200 ± 50 nM), the most potent inactivator of ICL1 yet characterized. Crystallographic and mass spectrometric analysis demonstrated that Cys191 of ICL1 was S-malylated by cis-EpS, and a crystallographic "snapshot" of inactivation lent insight into the chemical mechanism of this inactivation. Proteomic analysis of E. coli lysates showed that cis-EpS selectively labeled plasmid-expressed Mtb ICL1. Consistently, cis-EpS, but not its trans-isomer, inhibited the growth of Mtb under conditions in which ICL function is essential. These findings encourage the development of analogs of cis-2,3-epoxysuccinate as antituberculosis agents.
Asunto(s)
Antituberculosos/química , Inhibidores Enzimáticos/química , Isocitratoliasa/antagonistas & inhibidores , Mycobacterium tuberculosis/enzimología , Succinatos/química , Tuberculosis/tratamiento farmacológico , Antituberculosos/metabolismo , Descubrimiento de Drogas , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/metabolismo , Escherichia coli/metabolismo , Glicolatos/química , Glioxilatos/química , Humanos , Isomerismo , Modelos Moleculares , Unión Proteica , Conformación Proteica , Proteómica , Succinatos/metabolismo , TermodinámicaRESUMEN
Based on the specific and spontaneous formation of isopeptide bonds by SpyCatcher/SpyTag, we have developed a one-step method for purification and immobilization of recombinant proteins. The procedure is to immobilize SpyCatcher on glyoxyl agarose gels, and then the SpyCatcher immobilisate can be used to immobilize the SpyTag-fused protein in the crude extract selectively. A mutant of SpyCatcher (mSC), in which a peptide (LysGlyLysGlyLysGly) was added to the C-terminus of SpyCatcher and three lysine residues around the SpyTag/SpyCatcher binding domain were replaced with arginine, was designed to improve the attachment of SpyCatcher to the support. Compared with wild-type SpyCatcher, mSC can be immobilized on the glyoxyl-agarose support more efficiently, which enables the obtained mSC derivative a high binding capacity of the SpyTag-fused protein. The results showed that the target proteins in the crude enzyme extract were purified and immobilized in one step, and the thermal stability of the immobilized target proteins was also remarkably improved.
Asunto(s)
Proteínas Inmovilizadas/aislamiento & purificación , Proteínas Recombinantes/aislamiento & purificación , Adhesinas Bacterianas/química , Adhesinas Bacterianas/genética , Secuencia de Aminoácidos , Glioxilatos/química , Proteínas Inmovilizadas/química , Proteínas Inmovilizadas/metabolismo , Mutación , Oligopéptidos/química , Oligopéptidos/aislamiento & purificación , Estabilidad Proteica , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/aislamiento & purificación , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Sefarosa/química , TemperaturaRESUMEN
Cyclodextrins (CDs) and their derivatives have attracted significant attention in the pharmaceutical, food, and textile industries, which has led to an increased demand for their production. CD is typically produced by the action of cyclodextrin glycosyltransferase (CGTase) on starch. Owing to the relatively high cost of enzymes, the economic feasibility of the entire process strongly depends on the effective retention and recycling of CGTase in the reaction system, while maintaining its stability. CGTase enzymes immobilized on various supports such as porous glass beads or glyoxyl-agarose have been previously used to achieve this objective. Nevertheless, the attachment of biocatalysts on conventional supports is associated with numerous drawbacks, including enzyme leaching prominent in physical adsorption, reduced activity as a result of chemisorption, and increased mass transfer limitations. Recent reports on the successful utilization of metal-organic frameworks (MOFs) as supports for various enzymes suggest that CGTase could be immobilized for enhanced production of CDs. The three-dimensional microenvironment of MOFs could maintain the stability of CGTase while posing minimal diffusional limitations. Moreover, the presence of different functional groups on the surfaces of MOFs could provide multiple points for attachment of CGTase, thereby reducing enzyme loss through leaching. The present review focuses on the advantages MOFs can offer as support for CGTase immobilization as well as their potential for application in CD production.
Asunto(s)
Ciclodextrinas/química , Enzimas Inmovilizadas/química , Glucosiltransferasas/química , Estructuras Metalorgánicas/química , Glioxilatos/química , Sefarosa/química , Almidón/químicaRESUMEN
The reaction pathways were investigated by which a fungoid chitosan (CsG) may protect against photooxidative decay of model solutions and a sulphite-free white wine. Samples containing CsG were dark incubated for 2 days before exposure to fluorescent lighting for up to 21 days in the presence of wine like (+)-catechin and/or iron doses. In both systems CsG at winemaking doses significantly reduced the photoproduction of acetaldehyde and, to a better extent, glyoxylic acid, two key reactive aldehydes implicated in wine oxidative spoilage. After 21 days, CsG was two-fold more effective than sulphur dioxide in preventing glyoxylic acid formation and minimizing the browning of white wine. Among the antioxidant mechanisms involved in CsG protective effect, iron chelation, and hydrogen peroxide quenching were demonstrated. Besides, the previously unreported tartrate displacement from the [iron(III)-tartrate] complex was revealed as an additional inhibitory mechanism of CsG under photo-Fenton oxidation conditions.
Asunto(s)
Aldehídos/química , Quitosano/química , Procesos Fotoquímicos , Vino/análisis , Antioxidantes/química , Catequina/química , Glioxilatos/química , Oxidación-Reducción , Dióxido de Azufre/química , Tartratos/químicaRESUMEN
Deinococcus metallilatus MA1002 was exposed to ultraviolet radiation to generate mutants with enhanced biofilm production. Two strains (nos 5 and 6) were then selected based on their high biofilm formation, as well as their possession of higher concentrations of extracellular matrix components (eDNA, protein and saccharides) than the wild-type (WT). Genomic sequencing revealed the presence of large genome deletions in a secondary chromosome in the mutants. Expression analyses of the WT and mutant strains indicated the upregulation of genes associated with exopolysaccharide synthesis and stress response. The mutant strains showed high mortality in glucose-supplemented (TYG) medium; however, cell death and biofilm formation were not increased in mutant cells grown under acetate- or glyoxylate-added media, suggesting that metabolic toxicity during glucose metabolism induced a high rate of cell death but improved biofilm formation in mutant strains. In damaged cells, eDNAs contributed to the enhanced biofilm formation of D. metallilatus.
Asunto(s)
Proteínas Bacterianas/genética , Biopelículas/crecimiento & desarrollo , Deinococcus/crecimiento & desarrollo , Genómica/métodos , Mutación , Acetatos/química , Proteínas Bacterianas/efectos de la radiación , Biopelículas/efectos de la radiación , Medios de Cultivo/química , Deinococcus/genética , Deinococcus/efectos de la radiación , Evolución Molecular Dirigida , Perfilación de la Expresión Génica , Regulación Bacteriana de la Expresión Génica/efectos de la radiación , Glucosa/química , Glioxilatos/química , Viabilidad Microbiana , Análisis de Secuencia de ADN , Análisis de Secuencia de ARN , Secuenciación Completa del GenomaRESUMEN
The glyoxylate cycle is an important anabolic pathway and acts under a C2 compound (such as acetic acid) rich condition in bacteria. The isocitrate lyase (ICL) enzyme catalyzes the first step in the glyoxylate cycle, which is the cleavage of isocitrate to glyoxylate and succinate. This enzyme is a metalo-enzyme that contains an Mg2+ or a Mn2+ion at the active site for enzyme catalysis. We expressed and purified ICL from Bacillus cereus (BcICL) and investigated its biochemical properties and metal usage through its enzyme activity and stability with various divalent metal ion. Based on the results, BcICL mainly utilized the Mg2+ ion for enzyme catalysis as well as the Mn2+, Ni2+ and Co2+ ions. To elucidate its molecular mechanisms, we determined the crystal structure of BcICL at 1.79 Å. Through this structure, we analyzed a tetrameric interaction of the protein. We also determined the BcICL structure in complex with both the metal and its products, glyoxylate and succinate at 2.50 Å resolution and revealed each ligand binding modes.
Asunto(s)
Bacillus cereus/enzimología , Isocitratoliasa/química , Dominio Catalítico , Cristalografía por Rayos X , Glioxilatos/química , Isocitratoliasa/metabolismo , Magnesio/química , Metales/química , Modelos Moleculares , Multimerización de Proteína , Alineación de Secuencia , Análisis de Secuencia de Proteína , Ácido Succínico/químicaRESUMEN
Efforts to decipher the prebiotic roots of metabolic pathways have focused on recapitulating modern biological transformations, with metals typically serving in place of cofactors and enzymes. Here we show that the reaction of glyoxylate with pyruvate under mild aqueous conditions produces a series of α-ketoacid analogues of the reductive citric acid cycle without the need for metals or enzyme catalysts. The transformations proceed in the same sequence as the reverse Krebs cycle, resembling a protometabolic pathway, with glyoxylate acting as both the carbon source and reducing agent. Furthermore, the α-ketoacid analogues provide a natural route for the synthesis of amino acids by transamination with glycine, paralleling the extant metabolic mechanisms and obviating the need for metal-catalysed abiotic reductive aminations. This emerging sequence of prebiotic reactions could have set the stage for the advent of increasingly sophisticated pathways operating under catalytic control.
Asunto(s)
Ciclo del Ácido Cítrico/fisiología , Glioxilatos/química , Ácido Pirúvico/química , Catálisis , Ciclo del Ácido Cítrico/genética , Glioxilatos/metabolismo , Cetonas/química , Ácido Pirúvico/metabolismoRESUMEN
Oxidative stress, an excess of reactive oxygen species (ROS), may lead to oxidative post-translational modifications of proteins resulting in the cleavage of the peptide backbone, known as α-amidation, and formation of fragments such as peptide amides and α-ketoacyl peptides (α-KaP). In this study, we first compared different approaches for the synthesis of different model α-KaP and then investigated their stability compared to the corresponding unmodified peptides. The stability of peptides was studied at room temperature or at temperatures relevant for food processing (100 °C for cooking and 150 °C as a simulation of roasting) in water, in 1% (m/v) acetic acid or as the dry substance (to simulate the thermal treatment of dehydration processes) by HPLC analysis. Oxidation of peptides by 2,5-di-tert-butyl-1,4-benzoquinone (DTBBQ) proved to be the most suited method for synthesis of α-KaPs. The acyl side chain of the carbonyl-terminal α-keto acid has a crucial impact on the stability of α-KaPs. This carbonyl group has a catalytic effect on the hydrolysis of the neighboring peptide bond, leading to the release of α-keto acids. Unmodified peptides were significantly more stable than the corresponding α-KaPs. The possibility of further degradation reactions was shown by the formation of Schiff bases from glyoxylic or pyruvic acids with glycine and proven through detection of transamination products and Strecker aldehydes of α-keto acids by HPLC-MS/MS. We propose here a mechanism for the decomposition of α-ketoacyl peptides.
Asunto(s)
Péptidos/química , Aldehídos/química , Catálisis , Glicina/química , Glioxilatos/química , Hidrólisis , Cetoácidos/química , Oxidación-Reducción , Péptidos/síntesis química , Ácido Pirúvico/química , Bases de Schiff/químicaRESUMEN
Stabilization of dimeric enzymes requires the stabilization of the quaternary structure as well as the 3D one. Both subunits may be easily immobilized on a highly activated support. Additional stabilization of the 3D structure may be achieved via multipoint covalent attachment (MCA) on highly activated supports. In the case of monomeric enzymes or thermophilic dimeric ones, the optimal stabilization is obtained via the most intense MCA and it is associated to a small loss of catalytic activity. However, in the case of mesophilic enzymes, a very intense MCA of both subunits may promote negative effects, e.g., associated to distortions of the assembly between subunits and a subsequent very important loss of catalytic activity. A dimeric mesophilic amine oxidase from P.sativum was stabilized by MCA on glyoxyl-agarose. Both subunits were covalently immobilized on the support through the region with the highest density in Lys residues. In addition to that, an interesting activity/stabilization binomial was obtained after only 3 h of enzyme-support multiinteraction (50 % of activity/350 fold stabilization). However, after 24 h of enzyme-support multi-interaction this binomial activity-stabilization decreased down to 30/150. A moderate multiinteraction seems to be the optimal strategy for immobilization-stabilization of mesophilic dimeric enzymes and it promotes moderate losses of activity and interesting stabilizations against the combined effect of heat, acid pH and ethanol. The control of the intensity of enzyme-support multi-interactions becomes now strictly necessary.
Asunto(s)
Aminas , Enzimas Inmovilizadas/química , Oxidorreductasas/química , Pisum sativum/enzimología , Aminas/metabolismo , Estabilidad de Enzimas , Enzimas Inmovilizadas/metabolismo , Etanol/química , Glioxilatos/química , Concentración de Iones de Hidrógeno , Oxidorreductasas/metabolismo , Estructura Cuaternaria de Proteína , Subunidades de Proteína , Sefarosa/química , Temperatura , Factores de TiempoRESUMEN
The reactions of phenylglyoxylic acids during the synthesis and biological evaluation of fungal metabolites led to the discovery of hitherto unknown compounds with a p-quinone methide (p-QM) structure. The formation of these p-QMs using 13C-labelled starting materials revealed a key-step of this reaction being a retro-Friedel-Crafts alkylation.
Asunto(s)
Hongos , Glioxilatos/química , Ácidos Mandélicos/química , Hongos/química , Hongos/metabolismo , Glioxilatos/metabolismo , Espectroscopía de Resonancia Magnética , Ácidos Mandélicos/metabolismo , Modelos Moleculares , Conformación Molecular , Estructura Molecular , TemperaturaRESUMEN
The bi-enzymatic synthesis of the antiviral drug vidarabine (arabinosyladenine, ara-A), catalyzed by uridine phosphorylase from Clostridium perfringens (CpUP) and a purine nucleoside phosphorylase from Aeromonas hydrophila (AhPNP), was re-designed under continuous-flow conditions. Glyoxyl-agarose and EziGTM1 (Opal) were used as immobilization carriers for carrying out this preparative biotransformation. Upon setting-up reaction parameters (substrate concentration and molar ratio, temperature, pressure, residence time), 1 g of vidarabine was obtained in 55% isolated yield and >99% purity by simply running the flow reactor for 1 week and then collecting (by filtration) the nucleoside precipitated out of the exiting flow. Taking into account the substrate specificity of CpUP and AhPNP, the results obtained pave the way to the use of the CpUP/AhPNP-based bioreactor for the preparation of other purine nucleosides.
Asunto(s)
Antivirales/química , Enzimas Inmovilizadas/química , Purina-Nucleósido Fosforilasa/química , Vidarabina/química , Aeromonas hydrophila/enzimología , Biocatálisis , Reactores Biológicos , Biotransformación/efectos de los fármacos , Clostridium perfringens/enzimología , Enzimas Inmovilizadas/genética , Glioxilatos/química , Humanos , Ingeniería de Proteínas/métodos , Nucleósidos de Purina/química , Nucleósidos de Purina/metabolismo , Purina-Nucleósido Fosforilasa/genética , Sefarosa/química , Especificidad por Sustrato , Vidarabina/biosíntesis , Vidarabina/genéticaRESUMEN
A highly efficient quinine-derived primary-amine-catalyzed asymmetric aldol addition of hydroxyacetone to arylglyoxals is described. Structurally diverse anti-2,3-dihydroxy-1,4-diones were generated in high yields, with good diastereoselectivities and enantioselectivities.