Stereodivergent Protein Engineering of Fatty Acid Photodecarboxylase for Light‐Driven Kinetic Resolution of Sec‐Alcohol Oxalates

Angewandte Chemie International Edition, Journal Year: 2024, Volume and Issue: 63(10)

Published: Jan. 10, 2024

Stereodivergent engineering of one enzyme to create stereocomplementary variants for synthesizing optically pure molecules with tailor-made (R) or (S) configurations on an optional basis is highly desirable and challenging. This study aimed engineer fatty acid photodecarboxylase from Chlorella variabilis (CvFAP) using the focused rational iterative site-specific mutagenesis (FRISM) strategy obtain two excellent selectivity (both giving products up 99 % e.e.). These were used CvFAP-catalyzed light-driven kinetic resolution oxalates oxamic acids prepared corresponding sec-alcohols amines, providing a new biotransformation process preparing chiral amines. Molecular dynamics simulation, data transient spectra revealed source selectivity. represents first example amines catalyzed by pair CvFAPs.

Language: Английский

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An In Vitro Hybrid Biocatalytic System Enabled by a Combination of Surface-Displayed, Purified, and Cell-Free Expressed Enzymes

ACS Synthetic Biology, Journal Year: 2024, Volume and Issue: 13(5), P. 1434 - 1441

Published: May 2, 2024

Enzymatic cascades have become a green and sustainable approach for the synthesis of valuable chemicals pharmaceuticals. Using sequential enzymes to construct multienzyme complex is an effective way enhance overall performance biosynthetic routes. Here we report design efficient in vitro hybrid biocatalytic system by assembling three that can convert styrene (S)-1-phenyl-1,2-ethanediol. Specifically, prepared different ways, which were cell surface-displayed, purified, cell-free expressed. To assemble them, fused two orthogonal peptide–protein pairs (i.e., SpyTag/SpyCatcher SnoopTag/SnoopCatcher) enzymes, allowing their spatial organization covalent assembly. By doing this, constructed complex, could production (S)-1-phenyl-1,2-ethanediol 3 times compared free-floating enzyme without After optimization reaction system, final product yield reached 234.6 μM with substrate conversion rate 46.9% (based on 0.5 mM styrene). Taken together, our strategy integrates merits advanced biochemical engineering techniques, including cellular surface display, organization, expression, offers new solution chemical biosynthesis enzymatic cascade biotransformation. We, therefore, anticipate will hold great potential designing constructing highly systems synthesize agricultural, industrial, pharmaceutical significance.

Language: Английский

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Computational design of highly active de novo enzymes

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Aug. 3, 2024

Custom designed enzymes can further enhance the use of biocatalysts in industrial biotransformations, thereby helping to tackle biotechnological challenges 21st century. We present rotamer inverted fragment finder - diffusion (Riff-Diff) a hybrid machine learning and atomistic modeling strategy for scaffolding catalytic arrays de novo protein backbones with custom substrate pockets. used Riff-Diff scaffold tetrad capable efficiently catalyzing retro-aldol reaction. Functional designs exhibit high fold diversity, pockets similar natural enzymes. Some thus generated show activities rivaling those optimized by in-vitro evolution. The design can, principle, be applied any catalytically competent amino acid constellation. These findings are paving way address factors practical applicability catalysts processes shed light on fundamental principles enzyme catalysis.

Language: Английский

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A De Novo Metalloenzyme for Cerium Photoredox Catalysis

Journal of the American Chemical Society, Journal Year: 2024, Volume and Issue: 146(38), P. 25976 - 25985

Published: Aug. 8, 2024

Cerium photoredox catalysis has emerged as a powerful strategy to activate molecules under mild conditions. Radical intermediates are formed using visible light and simple complexes of the earth-abundant lanthanide. Here, we report an artificial photoenzyme enabling this chemistry inside protein. We utilize de novo designed protein scaffold that tightly binds lanthanide ions in its central cavity. Upon visible-light irradiation, cerium-dependent enzyme catalyzes radical C-C bond cleavage 1,2-diols aqueous solution. Protein engineering led variants with improved photostability metal binding behavior. The cleaves range aromatic aliphatic substrates, including lignin surrogates. Surface display on

Language: Английский

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FAIR Data and Software: Improving Efficiency and Quality of Biocatalytic Science

ACS Catalysis, Journal Year: 2024, Volume and Issue: 14(4), P. 2709 - 2718

Published: Feb. 7, 2024

Biocatalysis is entering a promising era as data-driven science. High-throughput experimentation generates rapidly increasing stream of biocatalytic data, which the raw material for mechanistic and modeling to design improved biocatalysts bioprocesses. However, our laboratory routines scientific practice communicating results are insufficient ensure reproducibility scalability experiments, data management has become bottleneck progress in biocatalysis. In order take full advantage rapid experimental computational technologies, should be findable, accessible, interoperable, reusable (FAIR). FAIRification software achieved by developing standardized exchange formats ontologies, electronic lab notebooks acquisition documentation experimentation, collaborative platforms analyzing repositories publishing together with data. The EnzymeML platform provides extensible tools FAIR scalable digitalization biocatalysis expected improve efficiency research automation guarantee quality science reproducibility. Most all, they foster reasoning creating hypotheses enabling reanalysis previously published thus promote disruptive innovation.

Language: Английский

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