- Electrochemical Activation of Galactose Oxidase: Mechanistic Studies and Synthetic Applications
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The enzyme galactose oxidase (GOase) is a copper radical oxidase that catalyzes the aerobic oxidation of primary alcohols to the aldehydes and has been utilized to that end in large-scale pharmaceutical processes. To maintain its catalytic activity and ensure high substrate conversion, GOase needs to be continuously (re)activated by 1e- oxidation of the constantly formed out-of-cycle species (GOasesemi) to the catalytically active state (GOaseox). In this work, we report an electrochemical activation method for GOase that replaces the previously used expensive horseradish peroxidase activator in a GOase-catalyzed oxidation reaction. First, the formation of GOaseox of a specifically engineered variant via nonenzymatic oxidation of GOasesemi was studied by UV-vis spectroscopy. Second, electrochemical oxidation of GOase by mediators was studied using cyclic voltammetry. The electron-transfer rates between GOase and various mediators at different pH values were determined, showing a dependence on both the redox potential of the mediator and the pH. This observation suggests that the oxidation of GOase by mediators at pH 7-9 likely occurs via a concerted proton-coupled electron-transfer (PCET) mechanism under anaerobic conditions. Finally, this electrochemical GOase activation method was successfully applied to the development of a bioelectrocatalytic GOase-mediated aerobic oxidation of benzyl alcohol derivatives, cinnamyl alcohol, and aliphatic polyols, including the desymmetrizing oxidation of 2-ethynylglycerol, a key step in the biocatalytic cascade used to prepare the promising HIV therapeutic islatravir.
- Fryszkowska, Anna,Klapars, Artis,Marshall, Nicholas,Ruccolo, Serge,Strotman, Neil A.,Zhang, Shaoguang
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p. 7270 - 7280
(2021/06/30)
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- Converging conversion - using promiscuous biocatalysts for the cell-free synthesis of chemicals from heterogeneous biomass
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Production of chemicals from lignocellulosic biomass has been proposed as a suitable replacement to petrochemicals. However, one inherent challenge of biomass utilization is the heterogeneity of the substrate resulting in the presence of mixed sugars after hydrolysis. Fermentation of mixed sugars often leads to poor yield and generation of multiple by-products, thus complicating the subsequent downstream processing. System biocatalysis has thus been developed in recent years to address this challenge. In this work, several novel enzymes with broad substrate promiscuity were identified using a sequence-based discovery approach as suitable biocatalysts in a conversion ofd-xylose andl-arabinose, two major constituents of hemicellulose found in plant biomass. These promiscuous enzymes enabled simultaneous biotransformation ofd-xylose andl-arabinose to yield 1,4-butanediol (BDO) with a maximum production rate of 3 g L?1h?1and a yield of >95%. This model system was further adapted toward the production of α-ketoglutarate (2-KG) from the pentoses using O2as a cosubstrate for cofactor recycling reaching a maximum production rate of 4.2 g L?1h?1and a yield of 99%. To verify the potential applicability of our system, we attempted to scale up the BDO and 2-KG production fromd-xylose andl-arabinose. Simple optimization and reaction engineering allowed us to obtain BDO and 2-KG titers of 18 g L?1and 42 g L?1, with theoretical yields of >75% and >99%, respectively. One of the promiscuous enzymes identified together with auxiliary promiscuous enzymes was also suitable for stereoconvergent synthesis from a mixture ofd-glucose andd-galactose, predominant sugars found in food waste streams and microalgae biomass.
- Pick, André,Sieber, Volker,Sutiono, Samuel
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supporting information
p. 3656 - 3663
(2021/06/06)
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- Prebiotic synthesis of 2-deoxy-d-ribose from interstellar building blocks promoted by amino esters or amino nitriles
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Understanding the prebiotic genesis of 2-deoxy-d-ribose, which forms the backbone of DNA, is of crucial importance to unravelling the origins of life, yet remains open to debate. Here we demonstrate that 20 mol% of proteinogenic amino esters promote the selective formation of 2-deoxy-d-ribose over 2-deoxy-d-threopentose in combined yields of ≥4%. We also demonstrate the first aldol reaction promoted by prebiotically-relevant proteinogenic amino nitriles (20 mol%) for the enantioselective synthesis of d-glyceraldehyde with 6% ee, and its subsequent conversion into 2-deoxy-d-ribose in yields of ≥ 5%. Finally, we explore the combination of these two steps in a one-pot process using 20 mol% of an amino ester or amino nitrile promoter. It is hence demonstrated that three interstellar starting materials, when mixed together with an appropriate promoter, can directly lead to the formation of a mixture of higher carbohydrates, including 2-deoxy-d-ribose.
- Steer, Andrew M.,Bia, Nicolas,Smith, David K.,Clarke, Paul A.
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supporting information
p. 10362 - 10365
(2017/09/25)
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- Attempts towards the synthesis of mupirocin-H
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The stereoselective synthesis of segments C1-C6 (3), C7-C12 (4) of mupirocin-H has been achieved. The synthetic procedure for the C1-C6 segment includes the zinc mediated allyl Grignard reaction with Rglyceraldehyde, Swern oxidation/Witting olefination reactions and followed by Sharpless asymmetric epoxidation. The C7-C12 segment was synthesized using again Sharpless asymmetric epoxidation on mono PMB protected 2-butene-1,4-diol, followed by regioselective opening of this epoxide with trimethyl aluminium. Both segments C1-C6 (3) and C7-C12 (4) possesses the five new stereogenic centers along with trans-olefin, but in various attempts condensation of 3 and 4 segments to give C-C bond forming parent segment (2) not affirmed, hence this work constitutes the synthesis of fragments C1-C6 (3) and C7-C12 (4) of mupirocin-H. {figure presented}
- Bommagani, Shobanbabu,Thodupunuri, Prashanth,Sharma, Gangavaram V.M.
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- METHOD FOR PRODUCING CARBOHYDRATES FROM DIHYDROXYACETONE
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The present invention relates to the use of dihydroxyacetone ("DHA") in the preparation of a number of natural and rare carbohydrates. The present invention comprises three stages. In the first stage of the present invention, syngas and formaldehyde are produced from natural gas, biogas, biomass and C02 from industrial plants including electricity generating plants, steel mills, cement factories and bio refineries. In the second stage of the present invention, formaldehyde and syngas from first stage are condensed to produce DHA. In the third stage of the present invention, DHA serves as a starting material for the synthesis of natural and rare carbohydrates using enzymes belonging to isomerase, aldolases, epimerase and transketolase groups.
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Paragraph 00152
(2016/12/26)
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- Governing chemistry of cellulose hydrolysis in supercritical water
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At extremely low reaction times (0.02 s), cellulose was hydrolyzed in supercritical water (T=400°C and P=25 MPa) to obtain a sugar yield higher than 95 wt %, whereas the 5-hydroxymethylfurfural (5-HMF) yield was lower than 0.01 wt %. If the reaction time was increased to 1 s, the main product was glycolaldehyde (60 wt %). Independently of the reaction time, the yield of 5-HMF was always lower than 0.01 wt %. To evaluate the reaction mechanism of biomass hydrolysis in pressurized water, several parameters (temperature, pressure, reaction time, and reaction medium) were studied for different biomasses (cellulose, glucose, fructose, and wheat bran). It was found that the H+ and OH- ion concentration in the reaction medium as a result of water dissociation is the determining factor in the selectivity. The reaction of glucose isomerization to fructose and the further dehydration to 5-HMF are highly dependent on the ion concentration. By an increase in the pOH/pH value, these reactions were minimized to allow control of 5-HMF production. Under these conditions, the retroaldol condensation pathway was enhanced, instead of the isomerization/dehydration pathway. Just add water: A reaction mechanism for cellulose hydrolysis that can explain the huge selectivity of biomass hydrolysis in supercritical water is presented. The model of the reaction mechanism has been validated by several experiments carried out in a continuous pilot plant capable at various conditions. It was found that the proton and hydroxide anion concentration in the medium due to water dissociation (represented by the ionic product of water, Kw) is the determining factor in the selectivity of the process.
- Cantero, Danilo A.,Bermejo, M. Dolores,Cocero, M. José
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p. 1026 - 1033
(2015/03/30)
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- Indium-mediated allenylation of aldehydes and its application in carbohydrate chemistry: Efficient synthesis of D -ribulose and 1-deoxy- D -ribulose
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A two-step reaction sequence starting with the indium-mediated allenylation of aldehydes with 4-bromo-2-butyn-1-ols and subsequent ozonolysis of the resulting allenylic product was developed to generate a variety of dihydroxyacetone derivatives. The regioselectivity of the indium-promoted C-C bond-forming reaction can be manipulated through hydroxy protecting groups on 4-bromo-2-butyn-1-ol, yielding either allenes or alkynes as preferred products. Compared to established protocols, the necessary amount of indium for this type of allenylation can be decreased by a factor of two to four. The versatility of this strategy was demonstrated in thestereoselective and straightforward synthesis of D-ribulose and 1-deoxy-D-ribulose. Copyright
- Fischer, Michael,Schmoelzer, Christoph,Nowikow, Christina,Schmid, Walther
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supporting information; experimental part
p. 1645 - 1651
(2011/05/05)
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- A likely possible origin of homochirality in amino acids and sugars on prebiotic earth
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For life to start on earth and elsewhere, it is critical that the building blocks - amino acids and sugars - be in predominant homochiral form. Over the past century, the origin of terrestrial prebiotic homochirality has been the subject of many speculations. In this Letter I summarize the experimental evidence for ways in which some meteoritic components could have led to the dominance of l amino acids and d sugars on earth, and the most likely way in which the original chiral excesses in the meteorites were formed.
- Breslow, Ronald
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supporting information; scheme or table
p. 2028 - 2032
(2011/05/09)
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- Borate as a phosphate ester mimic in aldolase-catalyzed reactions: Practical synthesis of L-fructose and L-iminocyclitols
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Dihydroxyacetone phosphate (DHAP)-dependent aldolases have been widely used for the organic synthesis of unnatural sugars or derivatives. The practicality of using DHAP-dependent aldolases is limited by their strict substrate specificity and the high cost and instability of DHAP. Here we report that the DHAP-dependent aldolase L-rhamnulose 1-phosphate aldolase (RhaD) accepts dihydroxyacetone (DHA) as a donor substrate in the presence of borate buffer, presumably by reversible in situ formation of DHA borate ester. The reaction appears to be irreversible, with the products thermodynamically trapped as borate complexes. We have applied this discovery to develop a practical one-step synthesis of the non-caloric sweetener L-fructose. L-Fructose was synthesized from racemic glyceraldehyde and DHA in the presence of RhaD and borate in 92% yield on a gram scale. We also synthesized a series of L-iminocyclitols, which are potential glycosidase inhibitors, in only two steps.
- Sugiyama, Masakazu,Hong, Zhangyong,Whalen, Lisa J.,Greenberg, William A.,Wong, Chi-Huey
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p. 2555 - 2559
(2007/10/03)
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- Engineering stereocontrol into an aldolase-catalysed reaction
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A novel thermostable aldolase has been developed for synthetic application, and substrate engineering has been used to induce stereocontrol into aldol reactions of this naturally-promiscuous enzyme.
- Lamble, Henry J.,Danson, Michael J.,Hough, David W.,Bull, Steven D.
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p. 124 - 126
(2007/10/03)
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- Structure-based mutagenesis approaches toward expanding the substrate specificity of D-2-deoxyribose-5-phosphate aldolase
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2-Deoxyribose-5-phosphate aldolase (DERA, EC 4.1.2.4) catalyzes the reversible aldol reaction between acetaldehyde and D-glyceraldehyde-3-phosphate to generate D-2-deoxyribose-5-phosphate. It is unique among the aldolases as it catalyzes the reversible asymmetric aldol addition reaction of two aldehydes. In order to expand the substrate scope and stereoselectivity of DERA, structure-based substrate design as well as site-specific mutation has been investigated. Using the 1.05 A crystal structure of DERA in complex with its natural substrate as a guide, five site-directed mutants were designed in order to improve its activity with the unnatural nonphosphorylated substrate, D-2-deoxyribose. Of these, the S238D variant exhibited a 2.5-fold improvement over the wild-type enzyme in the retroaldol reaction of 2-deoxyribose. Interestingly, this S238D mutant enzyme was shown to accept 3-azidopropinaldehyde as a substrate in a sequential asymmetric aldol reaction to form a deoxy-azidoethyl pyranose, which is a precursor to the corresponding lactone and the cholesterol-lowering agent Lipitor. This azidoaldehyde is not a substrate for the wild-type enzyme. Another structure-based design of new nonphosphorylated substrates was focused on the aldol reaction with inversion in enantioselectivity using the wild type or the S238D variant as the catalyst and 2-methyl-substituted aldehydes as substrates. An example was demonstrated in the asymmetric synthesis of a deoxypyranose as a new effective synthon for the total synthesis of epothilones. In addition, to facilitate the discovery of new enzymatic reactions, the engineered E. coli strain SELECT (Δace, adhC, DE3) was developed to be used in the future for selection of DERA variants with novel nonphosphorylated acceptor specificity.
- DeSantis, Grace,Liu, Junjie,Clark, David P.,Heine, Andreas,Wilson, Ian A.,Wong, Chi-Huey
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- One-pot synthesis of L-fructose using coupled multienzyme systems based on rhamnulose-1-phosphate aldolase
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Two methods have been developed for the highly efficient enzymatic synthesis of L-fructose: one is based on rhamnulose-1-phosphate aldolase and acid phosphatase using racemic glyceraldehyde and dihydroxyacetone phosphate as substrates; the other is to gen
- Franke, Dirk,Machajewski, Timothy,Hsu, Che-Chang,Wong, Chi-Huey
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p. 6828 - 6831
(2007/10/03)
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- Synthesis of carbohydrates via tandem use of the osmium-catalyzed asymmetric dihydroxylation and enzyme-catalyzed aldol addition reactions
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A new strategy is described for the asymmetric synthesis of carbohydrate derivatives via the tandem use of the osmium-catalyzed asymmetric dihydroxylation (AD) and aldolase-catalyzed aldol addition reactions. Both D- and L-forms of fructose, 6-deoxy-galacto-2-heptulose, and 6-phenyl-galacto-2-hexulose were synthesized to illustrate this methodology.
- Henderson, Ian,Sharpless, K. Barry,Wong, Chi-Huey
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p. 558 - 561
(2007/10/02)
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- Dispiroketals in synthesis: Preparation of a stable, sterically demanding glyceraldehyde ketal and diastereoselective reactions with simple organometallic reagents
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A preparation of the novel dispiroketal of D-glyceraldehyde is described. The diastereoselective addition of various organometallic reagents proceeds with anti selectivity due to the large steric bulk of the dispiroketal.
- Ley,Woods,Zanotti-Gerosa
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- Mechanism of formation of 2-furaldehyde from D-xylose
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The literature records two alternative hypotheses for the mechanism of dehydration of xylose to 2-furaldehyde (furfural), namely (1) a succession of reactions proceeding mainly via open-chain intermediates, and (2) an acid-catalyzed sequence proceeding through a 2,5-anhydride intermediate.The existing evidence for hypotheses (1) and (2) is reviewed and found to favor (2).The major products from xylose in water at 250 deg (with and without acid catalysis) have been investigated on a time-resolved basis.A kinetic model based on the second hypothesis is found to be consistent with the experimental data; whereas kinetic models based on the first hypothesis do not fit the data.
- Antal, Michael Jerry,Leesomboon, Tongchit,Mok, William S.,Richards, Geoffrey N.
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- Reversible and in Situ Formation of Organic Arsenates and Vanadates as Organic Phosphate Mimics in Enzymatic Reactions: Mechanistic Investigation of Aldol Reactions and Synthetic Applications
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A synthetic strategy is developed that uses organic phosphate utilizing enzymes as catalysts and a mixture of an organic alcohol and inorganic arsenate or vanadate to replace the organic phosphate substrate.In this process, inorganic arsenate or vanadate reacts with the alcohol reversibly in situ to form a mixture of esters, one of which is accepted by the enzyme as a substrate.Examples of the utility of this approach are demonstrated in enzymatic aldol condensations catalyzed by fructose-1,6-diphosphate aldolase, fuculose-1-phosphate aldolase, and rhamnulose-1-phosphate aldolase with a mixture of dihydroxyacetone and inorganic arsenate as substrate.Several uncommon sugars and deoxy sugars are prepared on 5-17-mmol scales.Mechanistic studies on an aldol reaction indicate that the redox reaction between dihydroxyacetone and inorganic vanadate prohibits the use of such a mixture to replace dihydroxyacetone phosphate in enzymatic aldol condensations.
- Drueckhammer, Dale G.,Durrwachter, J. Robert,Pederson, Richard L.,Crans, Debbie C.,Daniels, Lacy,Wong, Chi-Huey
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- Synthesis of Analogues of 1,3-Dihydroxyacetone Phosphate and Glyceraldehyde 3-Phosphate for Use in Studies of Fructose-1,6-diphosphate Aldolase
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This paper describes the synthesis of five analogues of dihydroxyacetone phosphate (3-azidohydroxyacetone 1-phosphate (5), 3-(acetylamino)hydroxyacetone 1-phosphate (12), (R)-1,3-dihydroxy-2-butanone 1-phosphate (18), (+/-)-1,3-dihydroxy-2-butanone 3-phosphate (26), and phosphonomethyl glycolate (31)).The syntheses of 18 and 26 are based on a new reaction: that is, the introduction of the phosphate group by the reaction of a diazo ketone with dibenzyl phosphate.These methods provide easy access to a number of compounds that are potential substrates for the synthetically useful enzyme aldolase (fructose-1,6-diphosphate aldolase from rabbit muscle, EC 4.1.2.13, RAMA) and perhaps for other enzymes of glycolysis.This paper also describes syntheses of 14 aldehydes for examination as substrates for aldolase.When the precursor was available, ozonolysis of vinyl groups proved to be the best route to the corresponding aldehydes.
- Bischofberger, Norbert,Waldmann, Herbert,Saito, Tohru,Simon, Ethan S.,Lees, Watson,et al.
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p. 3457 - 3465
(2007/10/02)
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- OPTICAL ACTIVITY OF D-GLYCERALDEHYDE IN AQUEOUS SOLUTIONS
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In dilute aqueous solutions, D-glyceraldehyde exists both in the free and hydrated monomeric forms; the equilibrium between them depends on the temperature.The specific rotation of the equilibrium mixture was measured polarimetrically in the temperature range from 20 to 80 deg C, the concentrations of both forms being determined by 1H NMR spectroscopy.From these data, the specific rotations of both forms of D-glyceraldehyde in the indicated temperature range were calculated.
- Fedoronko, Michal
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p. 1167 - 1172
(2007/10/02)
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- OXIDATIVE DEGRADATION OF MONOSACCHARIDES BY IRON(III) CHLORIDES WITH A NEAR-UV TO VISIBLE LIGHT UNDER AEROBIC ATMOSPHERE.
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Monosaccharides were oxidatively degraded by FeCl//3 with irradiation of a near-UV to visible light under aerobic atmosphere. D-Fructose and D-arabinose were degraded to D-erythrose in a yield of more than 80 mol%. D-Glucose and D-mannose were at first degraded to D-arabinose, which was subsequently degraded to D-erythrose, while d-ribose was directly degraded to d-erythrose and D-glyceraldehyde. The degradation processes were selective, and these reactions were shown to proceed via formation of an Fe(III)-monosaccharide complex. The kinetic data and other features of these reactions are also discussed.
- Araki,Sakuma,Shiraishi
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p. 997 - 1002
(2007/10/02)
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- Synthesis of Sugars by Aldolase-Catalyzed Condensation Reactions
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Dihydroxyacetone phosphate was prepared in 200-mmol scale from dihydroxyacetone by two procedures: reaction with phosphorus oxytrichloride and glycerol kinase catalyzed phosphorylation using ATP with in situ regeneration of ATP by phosphoenolpyruvate or acetyl phosphate.Dihydroxyacetone phosphate was converted to fructose 6-phosphate in 80percent yield by exposure to a mixture of co-immobilized triosephosphate isomerase and aldolase followed by acid hydrolysis of the condensation product fructose 1,6-bisphosphate.Fructose 6-phosphate was subsequently converted by chemical and enzymatic schemes into fructose, glucose 6-phosphate, and glucose.Practical procedures are described for the preparation of D- and L-glyceraldehyde 3-phosphate and for several hexoses labeled with 13C in the C-2 and C-2,5 positions.
- Wong, Chi-Huey,Whitesides, George M.
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p. 3199 - 3205
(2007/10/02)
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