Pub Date : 2023-05-24DOI: 10.3389/fctls.2023.1178345
Zoe Ingold, G. Grogan, Benjamin R. Lichman
Deoxypodophyllotoxin synthase (DPS) is a 2-oxoglutarate (2-OG) dependent non-heme iron (II) dioxygenase that catalyzes the stereoselective ring-closing carbon-carbon bond formation of deoxypodophyllotoxin from the aryllignan (−)-yatein. Deoxypodophyllotoxin is a precursor of topoisomerase II inhibitors, which are on the World Health Organization’s list of essential medicines. Previous work has shown that DPS can accept a range of substrates, indicating it has potential in biocatalytic processes for the formation of diverse polycyclic aryllignans. Recent X-ray structures of the enzyme reveal possible roles for amino acid side chains in substrate recognition and mechanism, although a mutational analysis of DPS was not performed. Here, we present a structure of DPS at an improved resolution of 1.41 Å, in complex with the buffer molecule, Tris, coordinated to the active site iron atom. The structure has informed a mutational analysis of DPS, which suggests a role for a D224-K187 salt bridge in maintaining substrate interactions and a catalytic role for H165, perhaps as the base for the proton abstraction at the final rearomatization step. This work improves our understanding of specific residues’ contributions to the DPS mechanism and can inform future engineering of the enzyme mechanism and substrate scope for the development of a versatile biocatalyst.
{"title":"Structure and mutation of deoxypodophyllotoxin synthase (DPS) from Podophyllum hexandrum","authors":"Zoe Ingold, G. Grogan, Benjamin R. Lichman","doi":"10.3389/fctls.2023.1178345","DOIUrl":"https://doi.org/10.3389/fctls.2023.1178345","url":null,"abstract":"Deoxypodophyllotoxin synthase (DPS) is a 2-oxoglutarate (2-OG) dependent non-heme iron (II) dioxygenase that catalyzes the stereoselective ring-closing carbon-carbon bond formation of deoxypodophyllotoxin from the aryllignan (−)-yatein. Deoxypodophyllotoxin is a precursor of topoisomerase II inhibitors, which are on the World Health Organization’s list of essential medicines. Previous work has shown that DPS can accept a range of substrates, indicating it has potential in biocatalytic processes for the formation of diverse polycyclic aryllignans. Recent X-ray structures of the enzyme reveal possible roles for amino acid side chains in substrate recognition and mechanism, although a mutational analysis of DPS was not performed. Here, we present a structure of DPS at an improved resolution of 1.41 Å, in complex with the buffer molecule, Tris, coordinated to the active site iron atom. The structure has informed a mutational analysis of DPS, which suggests a role for a D224-K187 salt bridge in maintaining substrate interactions and a catalytic role for H165, perhaps as the base for the proton abstraction at the final rearomatization step. This work improves our understanding of specific residues’ contributions to the DPS mechanism and can inform future engineering of the enzyme mechanism and substrate scope for the development of a versatile biocatalyst.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"42355792","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-15DOI: 10.3389/fctls.2023.1194977
Hakimeh Ardeshirfard, D. Elhamifar
In this study, magnetic cobalt oxide (Co3O4) nanoparticles (NPs) were synthesized through a new and green method using cobalt chloride hexahydrate (CoCl2.6H2O), pluronic P123 as a stabilizer, and sodium borohydride (NaBH4). The CO3O4 nanoparticles were characterized by diffuse reflectance infrared Fourier transform spectroscopy, powder X-ray diffraction, X-ray photoelectron spectroscopy, energy-dispersive X-ray spectroscopy, scanning electron microscopy, and vibrating sample magnetometer. The magnetic Co3O4 NPs were used as a catalyst with high activity and stability in the synthesis of tetrahydrobenzo[b]pyran derivatives. This reaction was carried out in water, as it is an environmentally friendly solvent, using a low loading of Co3O4 NPs at room temperature. Various derivatives of aldehydes were used as substrates to obtain a high yield of the corresponding tetrahydrobenzo[b]pyrans in short times. In addition, the catalyst was recovered and reused several times with no notable decrease in its activity.
{"title":"An efficient method for the preparation of magnetic Co3O4 nanoparticles and the study of their catalytic application","authors":"Hakimeh Ardeshirfard, D. Elhamifar","doi":"10.3389/fctls.2023.1194977","DOIUrl":"https://doi.org/10.3389/fctls.2023.1194977","url":null,"abstract":"In this study, magnetic cobalt oxide (Co3O4) nanoparticles (NPs) were synthesized through a new and green method using cobalt chloride hexahydrate (CoCl2.6H2O), pluronic P123 as a stabilizer, and sodium borohydride (NaBH4). The CO3O4 nanoparticles were characterized by diffuse reflectance infrared Fourier transform spectroscopy, powder X-ray diffraction, X-ray photoelectron spectroscopy, energy-dispersive X-ray spectroscopy, scanning electron microscopy, and vibrating sample magnetometer. The magnetic Co3O4 NPs were used as a catalyst with high activity and stability in the synthesis of tetrahydrobenzo[b]pyran derivatives. This reaction was carried out in water, as it is an environmentally friendly solvent, using a low loading of Co3O4 NPs at room temperature. Various derivatives of aldehydes were used as substrates to obtain a high yield of the corresponding tetrahydrobenzo[b]pyrans in short times. In addition, the catalyst was recovered and reused several times with no notable decrease in its activity.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"42659971","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-04DOI: 10.3389/fctls.2023.1147205
David Roura Padrosa, H. Lehmann, Radka Snajdrova, F. Paradisi
The application of phenylalanine ammonia lyases (PALs) for the amination of a variety of cinnamic acids has been shown to be a cost-efficient method to produce a variety of phenylalanine analogues. Nonetheless, as many other biocatalytic tools, the process intensification, especially due to the high equivalents of ammonia needed, and the cost-efficiency of the catalyst production and use have been key points to further prove their usefulness. Here, we investigated the use of previously characterized PALs (AvPAL and PbPAL) for the amination of a series of substituted cinnamic acids. To enhance the process scalability and the reusability of the catalyst, we investigated the use of covalent immobilization onto commercially available supports, creating a heterogeneous catalyst with good recovered activity (50%) and excellent stability. The immobilized enzyme was also incorporated in continuous flow for the synthesis of 3-methoxy-phenyl alanine and 4-nitro-phenylalanine, which allowed for shorter reaction times (20 min of contact time) and excellent conversions (88% ± 4% and 89% ± 5%) respectively, which could be maintained over extended period of time, up to 24 h. This work exemplifies the advantages that the combination of enzyme catalysis with flow technologies can have not only in the reaction kinetics, but also in the productivity, catalyst reusability and downstream processing.
{"title":"Sustainable synthesis of L-phenylalanine derivatives in continuous flow by immobilized phenylalanine ammonia lyase","authors":"David Roura Padrosa, H. Lehmann, Radka Snajdrova, F. Paradisi","doi":"10.3389/fctls.2023.1147205","DOIUrl":"https://doi.org/10.3389/fctls.2023.1147205","url":null,"abstract":"The application of phenylalanine ammonia lyases (PALs) for the amination of a variety of cinnamic acids has been shown to be a cost-efficient method to produce a variety of phenylalanine analogues. Nonetheless, as many other biocatalytic tools, the process intensification, especially due to the high equivalents of ammonia needed, and the cost-efficiency of the catalyst production and use have been key points to further prove their usefulness. Here, we investigated the use of previously characterized PALs (AvPAL and PbPAL) for the amination of a series of substituted cinnamic acids. To enhance the process scalability and the reusability of the catalyst, we investigated the use of covalent immobilization onto commercially available supports, creating a heterogeneous catalyst with good recovered activity (50%) and excellent stability. The immobilized enzyme was also incorporated in continuous flow for the synthesis of 3-methoxy-phenyl alanine and 4-nitro-phenylalanine, which allowed for shorter reaction times (20 min of contact time) and excellent conversions (88% ± 4% and 89% ± 5%) respectively, which could be maintained over extended period of time, up to 24 h. This work exemplifies the advantages that the combination of enzyme catalysis with flow technologies can have not only in the reaction kinetics, but also in the productivity, catalyst reusability and downstream processing.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-05-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"46899646","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-04-21DOI: 10.3389/fctls.2023.1154452
M. Crotti, M. S. Robescu, Juan M. Bolívar, D. Ubiali, L. Wilson, M. Contente
Flow biocatalysis is a key enabling technology that is increasingly being applied to a wide array of reactions with the aim of achieving process intensification, better control of biotransformations, and minimization of waste stream. In this mini-review, selected applications of flow biocatalysis to the preparation of food ingredients, APIs and fat- and oil-derived commodity chemicals, covering the period 2020-2022, are described.
{"title":"What’s new in flow biocatalysis? A snapshot of 2020–2022","authors":"M. Crotti, M. S. Robescu, Juan M. Bolívar, D. Ubiali, L. Wilson, M. Contente","doi":"10.3389/fctls.2023.1154452","DOIUrl":"https://doi.org/10.3389/fctls.2023.1154452","url":null,"abstract":"Flow biocatalysis is a key enabling technology that is increasingly being applied to a wide array of reactions with the aim of achieving process intensification, better control of biotransformations, and minimization of waste stream. In this mini-review, selected applications of flow biocatalysis to the preparation of food ingredients, APIs and fat- and oil-derived commodity chemicals, covering the period 2020-2022, are described.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"43345123","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-04-13DOI: 10.3389/fctls.2023.1165079
A. S. França, G. C. Brêda, K. D. de Oliveira, R. V. Almeida, F. Hollmann, R. D. de Souza
A visible-light-driven photocatalytic decarboxylation of palmitic acid and related fatty acids is described in this study. Remarkable decarboxylation rates have been observed with full conversion in less than 20 min. In this study, we have demonstrated that sunlight irradiation, even on cloudy days, can deliver similar results to traditional LED lamps while using much less energy and minimizing environmental impact. The findings indicate that the process of enzymatic decarboxylation could be useful for the production of different biofuels in the future.
{"title":"Impact of sunlight irradiation on CvFAP photodecarboxylation","authors":"A. S. França, G. C. Brêda, K. D. de Oliveira, R. V. Almeida, F. Hollmann, R. D. de Souza","doi":"10.3389/fctls.2023.1165079","DOIUrl":"https://doi.org/10.3389/fctls.2023.1165079","url":null,"abstract":"A visible-light-driven photocatalytic decarboxylation of palmitic acid and related fatty acids is described in this study. Remarkable decarboxylation rates have been observed with full conversion in less than 20 min. In this study, we have demonstrated that sunlight irradiation, even on cloudy days, can deliver similar results to traditional LED lamps while using much less energy and minimizing environmental impact. The findings indicate that the process of enzymatic decarboxylation could be useful for the production of different biofuels in the future.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-04-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"46325619","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-03-08DOI: 10.3389/fctls.2023.1114536
Sarah E. Cleary, Sofia Kazantzi, Joshua A. Trenchard, Martin Monedero, Jack W. Allman, Tara C. Lurshay, Xu Zhao, Michael B. C. Kenny, H. Reeve
Introduction: This work was carried out to understand if a heterogeneous biocatalytic hydrogenation system could be implemented in a slurry mode continuous flow reactor, as a “slot-in” alternative to a metal/carbon catalyst with minimal process or chemical engineering development. Method: Biocatalytic hydrogenation was compared to metal (Pd/C) catalysed hydrogenation for ketone to chiral alcohol conversion in both a continuous flow reactor (Coflore ACR, AM Technology) and analogous scaled down batch reactions. Results and discussion: Initial results demonstrated that batch reactions can achieve high conversions in 30 min, with relatively low biocatalyst loadings, meeting critical criteria for operation as continuous slurry mode process. Further results demonstrated full conversion of quinuclidinone to (3R)-quinuclidinol in continuous flow under mild conditions (35°C, 2 bar H2). On intensification of the process to higher substrate loading (50 mM), conversion was similar to with Pd/C, however the biocatalytic system achieved far higher turnover frequency and total turnover number (65 min−1 and 20,000, respectively) than the metal system (0.16 min−1, 37). Comparison to an analogous batch reaction highlights that the biocatalytic system has promise for further optimisation and intensification in the scalable Coflore ACR. Overall, biocatalytic hydrogenation is shown to offer a decarbonised biocatalytic route and a “slot-in” replacement to metal catalysts for hydrogenation reactions in continuous flow reactors.
{"title":"Preparation of (3R)-quinuclidinol using heterogeneous biocatalytic hydrogenation in a dynamically-mixed continuous flow reactor","authors":"Sarah E. Cleary, Sofia Kazantzi, Joshua A. Trenchard, Martin Monedero, Jack W. Allman, Tara C. Lurshay, Xu Zhao, Michael B. C. Kenny, H. Reeve","doi":"10.3389/fctls.2023.1114536","DOIUrl":"https://doi.org/10.3389/fctls.2023.1114536","url":null,"abstract":"Introduction: This work was carried out to understand if a heterogeneous biocatalytic hydrogenation system could be implemented in a slurry mode continuous flow reactor, as a “slot-in” alternative to a metal/carbon catalyst with minimal process or chemical engineering development. Method: Biocatalytic hydrogenation was compared to metal (Pd/C) catalysed hydrogenation for ketone to chiral alcohol conversion in both a continuous flow reactor (Coflore ACR, AM Technology) and analogous scaled down batch reactions. Results and discussion: Initial results demonstrated that batch reactions can achieve high conversions in 30 min, with relatively low biocatalyst loadings, meeting critical criteria for operation as continuous slurry mode process. Further results demonstrated full conversion of quinuclidinone to (3R)-quinuclidinol in continuous flow under mild conditions (35°C, 2 bar H2). On intensification of the process to higher substrate loading (50 mM), conversion was similar to with Pd/C, however the biocatalytic system achieved far higher turnover frequency and total turnover number (65 min−1 and 20,000, respectively) than the metal system (0.16 min−1, 37). Comparison to an analogous batch reaction highlights that the biocatalytic system has promise for further optimisation and intensification in the scalable Coflore ACR. Overall, biocatalytic hydrogenation is shown to offer a decarbonised biocatalytic route and a “slot-in” replacement to metal catalysts for hydrogenation reactions in continuous flow reactors.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-03-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"46449939","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-02-16DOI: 10.3389/fctls.2023.1116867
Hao Guo, Emily A. Gerstein, Kshitij C. Jha, Iskinder Arsano, M. Haider, T. Khan, M. Tsige
The present work quantifies metal-contaminant interactions between palladium substrates and three salient chlorinated organic contaminants, namely trichloroethylene 1,3,5-trichlorobenzene (TCB), and 3,3′,4,4′-tetrachlorobiphenyl (PCB77). Given that Pd is one of the conventional catalytically active materials known for contaminant removal, maximizing catalytic efficiency through optimal adsorption dynamics reduces the cost of remediation of contaminants that are persistent water pollutants chronically affecting public health. Adsorption efficiency analyses from all-atom molecular dynamics (MD) simulations advance the understanding of reaction mechanisms available from density functional theory (DFT) calculations to an extractable feature scale that can fit the parametric design of supported metal catalytic systems and feed into high throughput catalyst selection. Data on residence time, site-specific adsorption, binding energies, packing geometries, orientation profiles, and the effect of adsorbate size show the anomalous behaviour of organic contaminant adsorption on the undercoordinated {110} surface as compared to the {111} and {100} surfaces. The intermolecular interaction within contaminants from molecular dynamics simulation exhibits refreshing results than ordinary single molecule density functional theory calculation. Since complete adsorption and dechlorination is an essential step for chlorinated organic contaminant remediation pathways, the presented profiles provide essential information for designing efficient remediation systems through facet-controlled palladium nanoparticles. Graphical Abstract
{"title":"Non-reactive facet specific adsorption as a route to remediation of chlorinated organic contaminants","authors":"Hao Guo, Emily A. Gerstein, Kshitij C. Jha, Iskinder Arsano, M. Haider, T. Khan, M. Tsige","doi":"10.3389/fctls.2023.1116867","DOIUrl":"https://doi.org/10.3389/fctls.2023.1116867","url":null,"abstract":"The present work quantifies metal-contaminant interactions between palladium substrates and three salient chlorinated organic contaminants, namely trichloroethylene 1,3,5-trichlorobenzene (TCB), and 3,3′,4,4′-tetrachlorobiphenyl (PCB77). Given that Pd is one of the conventional catalytically active materials known for contaminant removal, maximizing catalytic efficiency through optimal adsorption dynamics reduces the cost of remediation of contaminants that are persistent water pollutants chronically affecting public health. Adsorption efficiency analyses from all-atom molecular dynamics (MD) simulations advance the understanding of reaction mechanisms available from density functional theory (DFT) calculations to an extractable feature scale that can fit the parametric design of supported metal catalytic systems and feed into high throughput catalyst selection. Data on residence time, site-specific adsorption, binding energies, packing geometries, orientation profiles, and the effect of adsorbate size show the anomalous behaviour of organic contaminant adsorption on the undercoordinated {110} surface as compared to the {111} and {100} surfaces. The intermolecular interaction within contaminants from molecular dynamics simulation exhibits refreshing results than ordinary single molecule density functional theory calculation. Since complete adsorption and dechlorination is an essential step for chlorinated organic contaminant remediation pathways, the presented profiles provide essential information for designing efficient remediation systems through facet-controlled palladium nanoparticles. Graphical Abstract","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-02-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49496176","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-02-16DOI: 10.3389/fctls.2023.1112154
C. N. Flores-Fernández, Max Cárdenas‐Fernández, G. Lye, John M. Ward
Co-expression of enzymes allow to produce multiple enzymes in a single host, representing a cost-effective alternative in biocatalytic processes which can be used for pectin bioconversion. Pectin-rich biomass is an abundant by-product from the fruit and sugar industries that is usually disposed in landfill or sold as a low value feedstock. The aim of this work was to co-express a thermophilic pectin methyl esterase (PME) and exo-polygalacturonases (exo-PGs) in a single host for pectin bioconversion into D-galacturonic acid (GalA) using different pectic substrates such as apple, citrus and sugar beet pectin. To achieve this, a PME from Bacillus licheniformis (BLI09) with either an exo-PG from Thermotoga maritima (TMA01) or from Bacillus licheniformis (BLI04) were cloned in pETDuet-1 and co-expressed in E. coli BL21 (DE3). Four co-expression plasmids containing both pectinases were constructed and factors such as the effect of the genes’ cloning order and their expression were evaluated. Co-expression constructs 3 and 4 (pETDuet-TMA01-BLI09 and pETDuet-BLI04-BLI09, respectively) showed better expression of both pectinases compared to co-expression constructs 1 and 2 (pETDuet-BLI09-TMA01 and pETDuet-BLI09-BLI04, respectively). Co-expression constructs 3 and 4 were the most efficient for pectin bioconversion into GalA reaching 3 and 2.5 mM GalA, respectively from apple and citrus pectin after 4 h reaction. In conclusion, this work demonstrates that the co-expression of pectinases can potentially contribute to reduce the cost associated to their production and purification as well as to increase their applicability for exploiting pectin-rich biomass to obtain bio-based chemicals.
酶的共表达允许在单个宿主中产生多种酶，这在可用于果胶生物转化的生物催化过程中是一种具有成本效益的替代方法。富含果胶的生物质是水果和糖业的大量副产品，通常被填埋或作为低价值原料出售。本工作的目的是在单一宿主中共同表达嗜热果胶甲基酯酶（PME）和外多聚半乳糖醛酸酶（exo-PGs），以使用不同的果胶底物如苹果、柑橘和甜菜果胶将果胶生物转化为D-半乳糖醛酸（GalA）。为了实现这一点，将来自地衣芽孢杆菌（BLI09）的PME与来自海洋嗜热菌（TMA01）或来自地衣芽孢菌（BLI04）的外显子PG克隆在pETDuet-1中，并在大肠杆菌BL21（DE3）中共表达。构建了四个含有两种果胶酶的共表达质粒，并对基因克隆顺序及其表达的影响等因素进行了评价。与共表达构建体1和2（分别为pETDuet-BLI09-TMA01和pETDue-BLI09-BLI04）相比，共表达构建物3和4（分别为p ETDuet-TMA01-BLI09和p ETDut-BLI04-BLI09）显示出更好的两种果胶酶表达。共表达构建体3和4对果胶生物转化为GalA最有效，在反应4小时后，从苹果和柑橘果胶分别达到3和2.5 mM GalA。总之，这项工作表明，果胶酶的共表达可能有助于降低其生产和纯化的相关成本，并提高其在利用富含果胶的生物质获得生物基化学品方面的适用性。
{"title":"Co-expression of thermophilic pectinases in a single host for cost-effective pectin bioconversion into D-galacturonic acid","authors":"C. N. Flores-Fernández, Max Cárdenas‐Fernández, G. Lye, John M. Ward","doi":"10.3389/fctls.2023.1112154","DOIUrl":"https://doi.org/10.3389/fctls.2023.1112154","url":null,"abstract":"Co-expression of enzymes allow to produce multiple enzymes in a single host, representing a cost-effective alternative in biocatalytic processes which can be used for pectin bioconversion. Pectin-rich biomass is an abundant by-product from the fruit and sugar industries that is usually disposed in landfill or sold as a low value feedstock. The aim of this work was to co-express a thermophilic pectin methyl esterase (PME) and exo-polygalacturonases (exo-PGs) in a single host for pectin bioconversion into D-galacturonic acid (GalA) using different pectic substrates such as apple, citrus and sugar beet pectin. To achieve this, a PME from Bacillus licheniformis (BLI09) with either an exo-PG from Thermotoga maritima (TMA01) or from Bacillus licheniformis (BLI04) were cloned in pETDuet-1 and co-expressed in E. coli BL21 (DE3). Four co-expression plasmids containing both pectinases were constructed and factors such as the effect of the genes’ cloning order and their expression were evaluated. Co-expression constructs 3 and 4 (pETDuet-TMA01-BLI09 and pETDuet-BLI04-BLI09, respectively) showed better expression of both pectinases compared to co-expression constructs 1 and 2 (pETDuet-BLI09-TMA01 and pETDuet-BLI09-BLI04, respectively). Co-expression constructs 3 and 4 were the most efficient for pectin bioconversion into GalA reaching 3 and 2.5 mM GalA, respectively from apple and citrus pectin after 4 h reaction. In conclusion, this work demonstrates that the co-expression of pectinases can potentially contribute to reduce the cost associated to their production and purification as well as to increase their applicability for exploiting pectin-rich biomass to obtain bio-based chemicals.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-02-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"46526695","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-02-03DOI: 10.3389/fctls.2022.1096824
Younes Abghoui, Atef Iqbal, Egill Skúlason
Following our previous report on N2 reduction reaction (NRR) on the surface of nitrides, we investigated the influence of incorporation of titanium nitride as a stable and inactive-NRR material into the structure of DFT-predicted NRR-active surfaces of chromium, vanadium, niobium, and zirconium nitrides. The outcome of our density functional theory (DFT) based analyses suggests that combination of titanium nitride with vanadium nitride can enhance the potential-determining step of the reaction with up to 20% compared to pure vanadium nitride while maintaining similar number of proton-electron transfer steps for formation of two ammonia molecules. The influence of titanium nitride on chromium nitride is expected to be more pronounced as rate-determining step associated with nitrogen adsorption on the vacancy and regeneration of the catalyst improves by around 90% compared to the pure chromium nitride. This effect on niobium and zirconium nitride is, however, negative as the potential-determining step becomes larger for the case of niobium nitride, and the reaction pathway changes from nitrogen reduction to hydrogen evolution for the case of zirconium nitride. These results not only encourage experimentalists to explore these overlayered structures further in experiments, but it also opens up the avenue for considering the alloys and dopants of these nitrides via both density functional theory modelling and experiments.
{"title":"The role of overlayered nitride electro-materials for N2 reduction to ammonia","authors":"Younes Abghoui, Atef Iqbal, Egill Skúlason","doi":"10.3389/fctls.2022.1096824","DOIUrl":"https://doi.org/10.3389/fctls.2022.1096824","url":null,"abstract":"Following our previous report on N2 reduction reaction (NRR) on the surface of nitrides, we investigated the influence of incorporation of titanium nitride as a stable and inactive-NRR material into the structure of DFT-predicted NRR-active surfaces of chromium, vanadium, niobium, and zirconium nitrides. The outcome of our density functional theory (DFT) based analyses suggests that combination of titanium nitride with vanadium nitride can enhance the potential-determining step of the reaction with up to 20% compared to pure vanadium nitride while maintaining similar number of proton-electron transfer steps for formation of two ammonia molecules. The influence of titanium nitride on chromium nitride is expected to be more pronounced as rate-determining step associated with nitrogen adsorption on the vacancy and regeneration of the catalyst improves by around 90% compared to the pure chromium nitride. This effect on niobium and zirconium nitride is, however, negative as the potential-determining step becomes larger for the case of niobium nitride, and the reaction pathway changes from nitrogen reduction to hydrogen evolution for the case of zirconium nitride. These results not only encourage experimentalists to explore these overlayered structures further in experiments, but it also opens up the avenue for considering the alloys and dopants of these nitrides via both density functional theory modelling and experiments.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":"2 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-02-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"43921216","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-01-16DOI: 10.3389/fctls.2023.1105948
Aurélie Fossey‐Jouenne, Laurine Ducrot, Ewald P. J. Jongkind, E. Elisée, A. Zaparucha, G. Grogan, Caroline E. Paul, C. Vergne‐Vaxelaire
Native amine dehydrogenases (nat-AmDHs) catalyze the (S)-stereoselective reductive amination of various ketones and aldehydes in the presence of high concentrations of ammonia. Based on the structure of CfusAmDH from Cystobacter fuscus complexed with Nicotinamide adenine dinucleotide phosphate (NADP+) and cyclohexylamine, we previously hypothesized a mechanism involving the attack at the electrophilic carbon of the carbonyl by ammonia followed by delivery of the hydride from the reduced nicotinamide cofactor on the re-face of the prochiral ketone. The direct reduction of carbonyl substrates into the corresponding alcohols requires a similar active site architecture and was previously reported as a minor side reaction of some native amine dehydrogenases and variants. Here we describe the ketoreductase (KRED) activity of a set of native amine dehydrogenases and variants, which proved to be significant in the absence of ammonia in the reaction medium but negligible in its presence. Conducting this study on a large set of substrates revealed the heterogeneity of this secondary ketoreductase activity, which was dependent upon the enzyme/substrate pairs considered. In silico docking experiments permitted the identification of some relationships between ketoreductase activity and the structural features of the enzymes. Kinetic studies of MsmeAmDH highlighted the superior performance of this native amine dehydrogenases as a ketoreductase but also its very low activity towards the reverse reaction of alcohol oxidation.
{"title":"Native amine dehydrogenases can catalyze the direct reduction of carbonyl compounds to alcohols in the absence of ammonia","authors":"Aurélie Fossey‐Jouenne, Laurine Ducrot, Ewald P. J. Jongkind, E. Elisée, A. Zaparucha, G. Grogan, Caroline E. Paul, C. Vergne‐Vaxelaire","doi":"10.3389/fctls.2023.1105948","DOIUrl":"https://doi.org/10.3389/fctls.2023.1105948","url":null,"abstract":"Native amine dehydrogenases (nat-AmDHs) catalyze the (S)-stereoselective reductive amination of various ketones and aldehydes in the presence of high concentrations of ammonia. Based on the structure of CfusAmDH from Cystobacter fuscus complexed with Nicotinamide adenine dinucleotide phosphate (NADP+) and cyclohexylamine, we previously hypothesized a mechanism involving the attack at the electrophilic carbon of the carbonyl by ammonia followed by delivery of the hydride from the reduced nicotinamide cofactor on the re-face of the prochiral ketone. The direct reduction of carbonyl substrates into the corresponding alcohols requires a similar active site architecture and was previously reported as a minor side reaction of some native amine dehydrogenases and variants. Here we describe the ketoreductase (KRED) activity of a set of native amine dehydrogenases and variants, which proved to be significant in the absence of ammonia in the reaction medium but negligible in its presence. Conducting this study on a large set of substrates revealed the heterogeneity of this secondary ketoreductase activity, which was dependent upon the enzyme/substrate pairs considered. In silico docking experiments permitted the identification of some relationships between ketoreductase activity and the structural features of the enzymes. Kinetic studies of MsmeAmDH highlighted the superior performance of this native amine dehydrogenases as a ketoreductase but also its very low activity towards the reverse reaction of alcohol oxidation.","PeriodicalId":73071,"journal":{"name":"Frontiers in catalysis","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2023-01-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"45078275","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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