@article{MTMT:34802673, title = {Transaminase-catalysis to produce trans-4-substituted cyclohexane-1-amines including a key intermediate towards cariprazine}, url = {https://m2.mtmt.hu/api/publication/34802673}, author = {Farkas, Emese and Sátorhelyi, Péter and Szakács, Zoltán and Dékány, Miklós and Vaskó, Dorottya and Hornyánszky, Gábor and Poppe, László and Éles, János}, doi = {10.1038/s42004-024-01148-9}, journal-iso = {COMMUN CHEM}, journal = {COMMUNICATIONS CHEMISTRY}, volume = {7}, unique-id = {34802673}, issn = {2399-3669}, abstract = {Cariprazine—the only single antipsychotic drug in the market which can handle all symptoms of bipolar I disorder—involves trans- 4-substituted cyclohexane-1-amine as a key structural element. In this work, production of trans -4-substituted cyclohexane-1-amines was investigated applying transaminases either in diastereotope selective amination starting from the corresponding ketone or in diastereomer selective deamination of their diasteromeric mixtures. Transaminases were identified enabling the conversion of the cis -diastereomer of four selected cis/trans -amines with different 4-substituents to the corresponding ketones. In the continuous-flow experiments aiming the cis diastereomer conversion to ketone, highly diastereopure trans -amine could be produced ( de > 99%). The yield of pure trans -isomers exceeding their original amount in the starting mixture could be explained by dynamic isomerization through ketone intermediates. The single transaminase-catalyzed process—exploiting the cis -diastereomer selectivity of the deamination and thermodynamic control favoring the trans -amines due to reversibility of the steps—allows enhancement of the productivity of industrial cariprazine synthesis.}, year = {2024}, eissn = {2399-3669}, orcid-numbers = {Szakács, Zoltán/0000-0001-6557-4409; Dékány, Miklós/0000-0001-8032-2998; Vaskó, Dorottya/0000-0002-2502-0644; Poppe, László/0000-0002-8358-1378} } @article{MTMT:34451217, title = {Upgrading Epoxy Supports for Enzyme Immobilization by Affinity Function Doping—A Case Study with Phenylalanine Ammonia-Lyase from Petroselinum crispum}, url = {https://m2.mtmt.hu/api/publication/34451217}, author = {Alács, Bálint and Zrinyi, Anna and Hornyánszky, Gábor and Poppe, László and Bell, Evelin}, doi = {10.3390/catal14010014}, journal-iso = {CATALYSTS}, journal = {CATALYSTS}, volume = {14}, unique-id = {34451217}, abstract = {This article provides a method to upgrade epoxy-functionalized carriers for covalent enzyme immobilization to selective carriers suitable for covalent immobilization of metal affinity-tagged enzymes without the need of preliminary enzyme purification. Affinity function doping of the epoxy-functionalized surface introduces an advanced possibility to avoid the costly and time-consuming downstream processes required for efficient immobilization on non-selective epoxy carriers. Our approach is based on the partial functionalization of surface epoxides via a proper diamine-derived linker and an ethylenediaminetetraacetic dianhydride-based chelator charged with cobalt ions. The solid macroporous carriers, doped with metal affinity functions, have both coordinative binding ability (rapid anchoring the metal affinity-tagged enzymes to the surface) and subsequent covalent bond-forming ability (preferred binding of the tagged enzyme to the surface after proper washing by the residual epoxide functions), enabling a single operation for the enrichment and immobilization of a recombinant phenylalanine ammonia-lyase from parsley fused to a polyhistidine affinity tag. The immobilized PcPAL was applied in the ammonia elimination of racemic phenylalanine, 4-chlorophenylalanine, and 4-bromophenylalanine to produce the corresponding d-phenylalanines, in addition to the formation of (E)-cinnamates, as well as in ammonia addition reactions to (E)-cinnamates, yielding the corresponding enantiopure l-phenylalanines.}, year = {2024}, eissn = {2073-4344}, orcid-numbers = {Poppe, László/0000-0002-8358-1378} } @article{MTMT:34138059, title = {Optimization Workflow of Fumonisin Esterase Production for Biocatalytic Degradation of Fumonisin B1}, url = {https://m2.mtmt.hu/api/publication/34138059}, author = {Incze, Dániel János and Poppe, László and Bata, Zsófia}, doi = {10.3390/life13091885}, journal-iso = {LIFE-BASEL}, journal = {LIFE-BASEL}, volume = {13}, unique-id = {34138059}, abstract = {Industrial enzyme production with the Pichia pastoris expression system requires a well-characterized production strain and a competitively priced fermentation medium to meet the expectations of the industry. The present work shows a workflow that allows the rapid and reliable screening of transformants of single copy insertion of the target production cassette. A constitutive expression system with the glyceraldehyde-3-phosphate dehydrogenase promoter (pGAP) with homology arms for the glycerol kinase 1 (GUT1) was constructed for the targeted integration of the expression plasmid in a KU70 deficient Pichia pastoris and the production of a bacterial fumonisin esterase enzyme (CFE). A robust colony qPCR method was developed for the copy number estimation of the expression cassette. Optimization of the protein production medium and the scale-up ability was aided by design of experiments (DOE) approach resulting in optimized production conditions at a semi-industrial scale. A novel fermentation medium containing 3% inactivated yeast and 2% dextrose in an ammonium-citrate buffer (IYD) was shown to be a promising alternative to YPD media (containing yeast extract, peptone, and dextrose), as similar protein titers could be obtained, while the cost of the medium was reduced 20-fold. In a demonstration-scale 48 h long fed-batch fermentation, the IYD media outperformed the small-scale YPD cultivation by 471.5 ± 22.6%.}, year = {2023}, eissn = {2075-1729}, orcid-numbers = {Poppe, László/0000-0002-8358-1378; Bata, Zsófia/0000-0002-4840-9433} } @article{MTMT:34063461, title = {Immobilization of Lipase B from Candida antarctica on Magnetic Nanoparticles Enhances Its Selectivity in Kinetic Resolutions of Chiral Amines with Several Acylating Agents}, url = {https://m2.mtmt.hu/api/publication/34063461}, author = {Silva, Fausto M. W. G. and Szemes, József and Mustashev, Akan and Takács, Orsolya and Imarah, Ali O. and Poppe, László}, doi = {10.3390/life13071560}, journal-iso = {LIFE-BASEL}, journal = {LIFE-BASEL}, volume = {13}, unique-id = {34063461}, abstract = {In lipase-catalyzed kinetic resolutions (KRs), the choice of immobilization support and acylating agents (AAs) is crucial. Lipase B from Candida antarctica immobilized onto magnetic nanoparticles (CaLB-MNPs) has been successfully used for diverse KRs of racemic compounds, but there is a lack of studies of the utilization of this potent biocatalyst in the KR of chiral amines, important pharmaceutical building blocks. Therefore, in this work, several racemic amines (heptane-2-amine, 1-methoxypropan-2-amine, 1-phenylethan-1-amine, and 4-phenylbutan-2-amine, (±)-1a–d, respectively) were studied in batch and continuous-flow mode utilizing different AAs, such as diisopropyl malonate 2A, isopropyl 2-cyanoacetate 2B, and isopropyl 2-ethoxyacetate 2C. The reactions performed with CaLB-MNPs were compared with Novozym 435 (N435) and the results in the literature. CaLB-MNPs were less active than N435, leading to lower conversion, but demonstrated a higher enantiomer selectivity, proving to be a good alternative to the commercial form. Compound 2C resulted in the best balance between conversion and enantiomer selectivity among the acylating agents. CaLB-MNPs proved to be efficient in the KR of chiral amines, having comparable or superior properties to other CaLB forms utilizing porous matrices for immobilization. An additional advantage of using CaLB-MNPs is that the purification and reuse processes are facilitated via magnetic retention/separation. In the continuous-flow mode, the usability and operational stability of CaLB-MNPs were reaffirmed, corroborating with previous studies, and the results overall improve our understanding of this potent biocatalyst and the convenient U-shape reactor used.}, year = {2023}, eissn = {2075-1729}, orcid-numbers = {Takács, Orsolya/0000-0002-4807-0396; Poppe, László/0000-0002-8358-1378} } @article{MTMT:33636410, title = {Scalability of U-Shape Magnetic Nanoparticles-Based Microreactor–Lipase-Catalyzed Preparative Scale Kinetic Resolutions of Drug-like Fragments}, url = {https://m2.mtmt.hu/api/publication/33636410}, author = {Silva, Fausto M. W. G. and Imarah, Ali O. and Takács, Orsolya and Tuba, László and Poppe, László}, doi = {10.3390/catal13020384}, journal-iso = {CATALYSTS}, journal = {CATALYSTS}, volume = {13}, unique-id = {33636410}, abstract = {The production of active pharmaceutical ingredients (APIs) and fine chemicals is accelerating due to the advent of novel microreactors and new materials for immobilizing customized biocatalysts that permit long-term use in continuous-flow reactors. This work studied the scalability of a tunable U-shape magnetic nanoparticles (MNPs)-based microreactor. The reactor consisted of a polytetrafluoroethylene tube (PTFE) of various inner diameters (ID = 0.75 mm, 1.50 mm, or 2.15 mm) and six movable permanent magnets positioned under the tube to create reaction chambers allowing the fluid reaction mixture to flow through and above the enzyme-loaded MNPs anchored by permanent magnets. The microreactors with various tube sizes and MNP capacities were tested with the preparative scale kinetic resolution of the drug-like alcohols 4-(3,4-dihydroisoquinolin-2(1H)-yl)butan-2-ol (±)-1a and 4-(3,4-dihydroquinolin-1(2H)-yl)butan-2-ol (±)-1b, utilizing Lipase B from Candida antarctica immobilized covalently onto MNPs, leading to highly enantioenriched products [(R)-2a,b and (S)-1a,b]. The results in the U-shape MNP flow reactor were compared with reactions in the batch mode with CaLB-MNPs using similar conditions. Of the three different systems, the one with ID = 1.50 mm showed the best balance between the maximum loading capacity of biocatalysts in the reactor and the most effective cross-section area. The results showed that this U-shaped tubular microreactor might be a simple and flexible instrument for many processes in biocatalysis, providing an easy-to-set-up alternative to existing techniques.}, year = {2023}, eissn = {2073-4344}, pages = {384}, orcid-numbers = {Takács, Orsolya/0000-0002-4807-0396; Poppe, László/0000-0002-8358-1378} } @article{MTMT:33620537, title = {Magnetically agitated continuous-flow tube reactors with aspartate ammonia-lyase immobilized on magnetic nanoparticles}, url = {https://m2.mtmt.hu/api/publication/33620537}, author = {Imarah, Ali O. and Silva, Fausto M. W. G. and Bataa, Naran and Decsi, Balázs and Balogh Weiser, Diána and Poppe, László}, doi = {10.1039/D2RE00507G}, journal-iso = {REACT CHEM ENG}, journal = {REACTION CHEMISTRY & ENGINEERING}, volume = {8}, unique-id = {33620537}, issn = {2058-9883}, abstract = {Two magnetically agitated continuous-flow tube reactors (AFRs)—applying external permanent magnets to move magnetic nanoparticles coated with a biocatalyst within a stream of the reaction medium—were developed and studied with aspartate ammonia-lyase (AAL) from Pseudomonas fluorescens immobilized onto epoxy-functionalized magnetic nanoparticles (MNPs) by covalent binding. The biotransformation of L-aspartate to fumarate by the AAL-MNPs (5 mg, D = 420 nm, 6 μg g−1 AAL) took place in the reaction tube (PTFE, ID 2.15 mm) of both AFRs in a space containing the AAL-MNPs agitated within the flow of the reaction medium (0.5 mM L-aspartate solution, 4.7–14 μL min−1, 25 °C) by two permanent ring magnets (N48 neodymium, 10 × 5 × 5 mm) positioned at a fixed distance in attraction mode. In the first version (AFRXM), the two magnets positioned at opposite sides of the reaction tube (distance: 20 mm) performed axial movement (amplitude: 8 mm, frequency: 40–140 mpm) along the Y-axis, being perpendicular to the X-axis of the tube. In the second version (AFRRM), the two magnets (distance: 10 mm from each other, 5 mm from the X-axis) performed rotation movement (frequency: 40–140 rpm) around the X-axis. Whereas in the AFRXM the AAL-MNPs formed a cloud moving back and forth, they created a ring-shaped cloud rotating within the tube in the AFRRM. The efficient internal mixing in the AFRRM at the best frequency (80 rpm) resulted in the highest apparent specific activity (Ub = 354–469 U g−1, at residence times of 2.5–7.5 min) of the AAL-MNPs in the reactors studied. In the other continuous-flow systems, significantly lower Ub values were achieved (135–290 U g−1 at 120 mpm in AFRXM; or 142–273 U g−1 and 64–129 U g−1 in tubular reactors anchoring MNPs in static mode with double or single magnets, respectively), whereas more than a magnitude of order lower values could be realised in the batch mode reactors (11.4–14.9 U g−1 with rotational magnetic agitation at 120 rpm; 5.0–5.8 U g−1 with axial magnetic agitation at 160 mpm; or 4.6–5.2 U g−1 in an orbital shaker at 600 rpm) at comparable reaction times (2.5–7.5 min).}, year = {2023}, eissn = {2058-9883}, pages = {1250-1259}, orcid-numbers = {Bataa, Naran/0000-0001-8827-250X; Balogh Weiser, Diána/0000-0002-9957-1203; Poppe, László/0000-0002-8358-1378} } @article{MTMT:33574611, title = {Novel Approach for the Isolation and Immobilization of a Recombinant Transaminase. Applying an Advanced Nanocomposite System}, url = {https://m2.mtmt.hu/api/publication/33574611}, author = {Koplányi, Gábor and Bell, Evelin and Molnár, Zsófia Klára and Katona, Gábor and Neumann, Péter Lajos and Ender, Ferenc and Balogh, György Tibor and Žnidaršič-Plazl, Polona and Poppe, László and Balogh Weiser, Diána}, doi = {10.1002/cbic.202200713}, journal-iso = {CHEMBIOCHEM}, journal = {CHEMBIOCHEM}, volume = {24}, unique-id = {33574611}, issn = {1439-4227}, year = {2023}, eissn = {1439-7633}, orcid-numbers = {Koplányi, Gábor/0000-0002-3791-1057; Katona, Gábor/0000-0003-1564-4813; Neumann, Péter Lajos/0000-0002-2881-5733; Ender, Ferenc/0000-0003-3800-5707; Balogh, György Tibor/0000-0003-3347-1880; Poppe, László/0000-0002-8358-1378; Balogh Weiser, Diána/0000-0002-9957-1203} } @article{MTMT:33633153, title = {Production of recombinant, non-tagged phenylalanine ammonia-lyases employing tev protease-removable affinity tags}, url = {https://m2.mtmt.hu/api/publication/33633153}, author = {Filip, Alina and Bata, Zsófia and Anghel, Anca Elena and Poppe, László and László Csaba, Bencze}, doi = {10.24193/subbchem.2022.4.03}, journal-iso = {STUD UNIV BABES-BOLYAI CHEM}, journal = {STUDIA UNIVERSITATIS BABES-BOLYAI CHEMIA}, volume = {67}, unique-id = {33633153}, issn = {1224-7154}, year = {2022}, eissn = {2065-9520}, pages = {27-46}, orcid-numbers = {Bata, Zsófia/0000-0002-4840-9433; Poppe, László/0000-0002-8358-1378} } @article{MTMT:33229099, title = {Novel biomimetic nanocomposite for investigation of drug metabolism}, url = {https://m2.mtmt.hu/api/publication/33229099}, author = {Balogh Weiser, Diána and Poppe, László and Kenéz, Balázs and Decsi, Balázs and Koplányi, Gábor and Katona, Gábor and Gyarmati, Benjámin Sándor and Ender, Ferenc and Balogh, György Tibor}, doi = {10.1016/j.molliq.2022.120781}, journal-iso = {J MOL LIQ}, journal = {JOURNAL OF MOLECULAR LIQUIDS}, volume = {368}, unique-id = {33229099}, issn = {0167-7322}, abstract = {In vitro mimicking of hepatic drug metabolism is a key issue in early-stage drug discovery. Synthetic metalloporphyrins show structural similarity with the heme type prosthetic group of cytochrome P450 as primary hepatic enzyme in oxidative drug biotransformation. Therefore, they can catalyze these oxidations. Concerning economical aspects and the poor stability of metalloporphyrin, their immobilization onto or into solid carriers can be promising solution. This study presents a novel immobilized metalloporphyrin nanocomposite system and its potential use as biomimetic catalysts. The developed two-step immobilization procedure consists of two main steps. First, the ionic binding of meso-tetra (parasulphonatophenyl) iron porphyrin onto functionalized magnetic nanoparticles is established, followed by embedding the nanoparticles into polylactic acid nanofibers by electrospinning technique. Due to the synergistic morphological and chemo-structural advantages of binding onto nanoparticles and embedding in polymeric matrices the biomimetic efficiency of metalloporphyrin can be remarkably enhanced, while substrate conversion value was tenfold larger than which could be achieved with classic human liver microsomal system.}, keywords = {IMMOBILIZATION; COMPOSITES; OXIDATION; NANOFIBERS; Biotransformation; Chemistry, Physical; FABRICATION; magnetic nanoparticles; Metalloporphyrins; CATALYTIC-ACTIVITY}, year = {2022}, eissn = {1873-3166}, orcid-numbers = {Balogh Weiser, Diána/0000-0002-9957-1203; Poppe, László/0000-0002-8358-1378; Koplányi, Gábor/0000-0002-3791-1057; Katona, Gábor/0000-0003-1564-4813; Ender, Ferenc/0000-0003-3800-5707; Balogh, György Tibor/0000-0003-3347-1880} } @article{MTMT:33099063, title = {A Convenient U-Shape Microreactor for Continuous Flow Biocatalysis with Enzyme-Coated Magnetic Nanoparticles–Lipase-Catalyzed Enantiomer Selective Acylation of 4-(Morpholin-4-yl)butan-2-ol}, url = {https://m2.mtmt.hu/api/publication/33099063}, author = {Imarah, Ali O. and Silva, Fausto M. W. G. and Tuba, László and Malta-Lakó, Ágnes and Szemes, József and Bell, Evelin and Poppe, László}, doi = {10.3390/catal12091065}, journal-iso = {CATALYSTS}, journal = {CATALYSTS}, volume = {12}, unique-id = {33099063}, abstract = {This study implements a convenient microreactor for biocatalysis with enzymes immobilized on magnetic nanoparticles (MNPs). The enzyme immobilized onto MNPs by adsorption or by covalent bonds was lipase B from Candida antarctica (CaLB). The MNPs for adsorption were obtained by covering the magnetite core with a silica shell and later with hexadecyltrimethoxysilane, while for covalent immobilization, the silica-covered MNPs were functionalized by a layer forming from mixtures of hexadecyl- and 3-(2-aminoethylamino)propyldimethoxymethylsilanes in 16:1 molar ratio, which was further activated with neopentyl glycol diglycidyl ether (NGDE). The resulting CaLB-MNPs were tested in a convenient continuous flow system, created by 3D printing to hold six adjustable permanent magnets beneath a polytetrafluoroethylene tube (PTFE) to anchor the MNP biocatalyst inside the tube reactor. The anchored CaLB-MNPs formed reaction chambers in the tube for passing the fluid through and above the MNP biocatalysts, thus increasing the mixing during the fluid flow and resulting in enhanced activity of CaLB on MNPs. The enantiomer selective acylation of 4-(morpholin-4-yl)butan-2-ol (±)-1, being the chiral alcohol constituent of the mucolytic drug Fedrilate, was carried out by CaLB-MNPs in the U-shape reactor. The CaLB-MNPs in the U-shape reactor were compared in batch reactions to the lyophilized CaLB and to the CaLB-MNPs using the same reaction composition, and the same amounts of CaLB showed similar or higher activity in flow mode and superior activity as compared to the lyophilized powder form. The U-shape permanent magnet design represents a general and easy-to-access implementation of MNP-based flow microreactors, being useful for many biotransformations and reducing costly and time-consuming downstream processes.}, year = {2022}, eissn = {2073-4344}, orcid-numbers = {Malta-Lakó, Ágnes/0000-0002-9449-0957; Poppe, László/0000-0002-8358-1378} }