Synthesis of 1-cyanocyclohexaneacetic acid in a partial-mixed recirculating bed reactor with resin-immobilized nitrilase.

Immobilized enzyme bioreactors provide a controlled reaction environment and integrated reaction-separation processes for biocatalysis. In this study, a biocatalytic process based on resin-immobilized nitrilase and a partial-mixed recirculating bed bioreactor was developed for the synthesis of 1-cyanocyclohexaneacetic acid, a gabapentin intermediate. The resin catalyst prepared by immobilizing the regioselective nitrilase AcN-S on the activated amino resin LX-1000EPHA achieved 99.

Restore the Activity and Stability of Nicotinamide Riboside Kinase by α-Helix Reconstruction and Semi-Rational Design.

Nicotinamide riboside kinase (NRK) is an important enzyme in the nicotinamide riboside (NR) metabolic pathway, converting NR to nicotinamide mononucleotide (NMN), which has promising industrial applications. However, structural collapse in many NRKs reduces their activity and thermal stability, limiting their industrial potential. Herein, we developed an α-helix reconstruction method on a Kag-NRK from Kluyveromyces marxianus for restore its activity and stability together.

Identification of a novel D-amino acid oxidase and its application in deracemization of D, L-phosphinothricin.

DAAO is applied as a potential catalyst in the biosynthesis of L-PPT. However, its low solubility expression constrains its broader industrial application. Herein, a novel DAAO derived from Cladophialophora carrionii (CcDAAO) was identified, which demonstrated superior catalytic performance toward D-Ala (specific activity: 106.

Characterization of the G protein-coupled receptor FfGpr1-Gα-AC signal transduction in Fusarium fujikuroi and its role in gibberellin biosynthesis.

Aims: This study aimed to characterize the G protein-coupled receptor FfGpr1-Gα-AC transduction pathway and its role in regulating gibberellin (GA) metabolism in Fusarium fujikuroi.

Methods And Results: By constructing Ffgpr1 deletion and constitutively activated FfG2Q204 L mutants, we found that glucose-induced cAMP (cyclic Adenosine MonoPhosphate) synthesis was abolished in Ffgpr1Δ. Bimolecular fluorescence complementation confirmed membrane-localized FfGpr1-FfG2 and FfG2-AC interactions, with intensified fluorescence at septa.

Design of a Distal Site Saturation Test-Iterative Parallel Mutagenesis for Engineering Hydroxysteroid Dehydrogenase.

Distal protein engineering facilitates the efficient identification of novel modification sites and synergistic modulation of enzyme functions to meet the demands of biocatalysts for industrial applications. Using hydroxysteroid dehydrogenase as a target protein, this study presents a distal site saturation test-iterative parallel mutagenesis (DSST-IPM) strategy to design high-performance enzymes. Twelve single-point mutations were identified to improve the stability-activity trade-off in the distal site, targeting 34 residues.

Resolving the Trade-Off Between Toxicity and Efficiency of CRISPR-Cas9 System for Genome Editing Within Escherichia coli.

Efficient gene editing of Escherichia coli BL21 (DE3) holds significant practical value as a host for heterologous protein expression. Recently reported CRISPR-Cas9 editing systems for this strain exhibit a trade-off between efficiency and toxicity. In this study, we addressed this trade-off by employing the strategy to transiently induce Cas9 expression in the high-copy plasmid during the editing stage.

The pedal-like loop of (R)-selective transaminases plays a critical role to the functionality of the enzyme.

In enzyme engineering, a lot of studies have focused on engineering the active site to broaden substrate specificity or enhance transaminase activity; however, relatively little is known about the mechanisms by which substrates are recognized and enter the binding pocket. Transaminases play a crucial role in the synthesis of chiral amines due to their exceptional stereoselectivity and catalytic efficiency. In this study, we explored how the pedal-like loop at the active site influences (R)-transaminase (ATA) activity and substrate recognition by modulating the substrate channel.

Advances in biosynthesis of chiral amide herbicides and the key enzymes: dimethenamid-P and S-metolachlor as case studies.

Chiral amide herbicides represent a significant class of agrochemicals, widely used for effective weed control. Prominent examples include S-metolachlor and dimethenamid-P, both of which share the intermediate (S)-1-methoxy-2-propylamine, a key structural component in their synthesis. Developing green and sustainable methods for producing this intermediate is crucial for enhancing the environmental and economic feasibility of herbicide manufacturing.

Nitrogen removal characteristics and salt tolerance mechanisms of the novel bacterium Halomonas sp. W07 in saline wastewater treatment.

The extremely high osmotic pressure that frequently emerges in industrial wastewater will notably impact microorganisms' survival and nitrogen removal efficiency. A newly isolated Halomonas sp. strain W07 demonstrated the ability to efficiently remove nitrate and nitrite at an average rate of 4.

Salt-driven dynamic folding of halophile-origin enzymes: Insights into evolution and protein exploitation.

Many years ago, life transitioned from the ocean to land, evolving from halophilic to non-halophilic organisms. Our research indicates that some enzymes from halophiles require salt for soluble expression in E. coli and retain activity within certain salt concentration ranges in the growth medium.

Avermectins and Their Derivatives: Recent Advances in Biosynthesis and Application.

Avermectins (AVMs) and their derivatives are the most effective and widely used nematicides, insecticides, and acaricides against endo- and ectoparasites of plants, animals, and humans. Demand for avermectins and their highly effective derivatives has increased due to their high cost-effectiveness and wide range of applications as medicines and crop protection products. Due to the unique structures of these compounds and for industrial production purposes, numerous efforts and strategies have been dedicated to enhancing the production of avermectins and creating new analogues in recent years.

Assembly and engineering of BioBricks to develop an efficient NADH regeneration system.

Unlabelled: The cofactor regeneration system plays a crucial role in redox biocatalysis for organic synthesis and the pharmaceutical industry. The alcohol dehydrogenase (ADH)-based regeneration system offers a promising solution for the regeneration of NAD(P)H. However, its widespread use is hindered by low activity and poor expression of ADH in .

Engineering of a hydroxysteroid dehydrogenase with simultaneous enhancement in activity and thermostability for efficient biosynthesis of ursodeoxycholic acid.

Unlabelled: Hydroxysteroid dehydrogenases (HSDHs) catalyze the oxidation/reduction of hydroxyl/keto groups of steroids with high regio- or stereoselectivity, playing an essential role in producing optically pure chemicals. In this work, a novel approach was developed to simultaneously improve the stability and activity of 7β-hydroxysteroid dehydrogenase (7β-HSDH) by combining B-factor analysis and computer-aided prediction. Several advantageous mutants were identified, and the most promising variant, S51Y/P202Y, exhibited 2.

Design of NAMPTs with Superior Activity by Dual-Channel Protein Engineering Strategy.

The nicotinamide phosphoribosyltransferase (NAMPT)-catalyzed substitution reaction plays a pivotal role in the biosynthesis of nucleotide compounds. However, industrial applications are hindered by the low activity of NAMPTs. In this study, a novel dual-channel protein engineering strategy was developed to increase NAMPT activity by enhancing substrate accessibility.

Development of growth selection system and pocket engineering of d-amino acid oxidase to enhance selective deamination activity toward d-phosphinothricin.

D-amino acid oxidase (DAAO)-catalyzed selective oxidative deamination is a very promising process for synthesizing l-amino acids including l-phosphinothricin (l-PPT, a high-efficiency and broad-spectrum herbicide). However, the wild-type DAAO's low activity toward unnatural substrates like d-phosphinothricin (d-PPT) hampers its application. Herein, a DAAO from Caenorhabditis elegans (CeDAAO) was screened and engineered to improve the catalytic potential on d-PPT.

Enhancing the expression of the unspecific peroxygenase in Komagataella phaffii through a combination strategy.

The unspecific peroxygenase (UPO) from Cyclocybe aegerita (AaeUPO) can selectively oxidize C-H bonds using hydrogen peroxide as an oxygen donor without cofactors, which has drawn significant industrial attention. Many studies have made efforts to enhance the overall activity of AaeUPO expressed in Komagataella phaffii by employing strategies such as enzyme-directed evolution, utilizing appropriate promoters, and screening secretion peptides. Building upon these previous studies, the objective of this study was to further enhance the expression of a mutant of AaeUPO with improved activity (PaDa-I) by increasing the gene copy number, co-expressing chaperones, and optimizing culture conditions.

Maximizing the potential of nitrilase: Unveiling their diversity, catalytic proficiency, and versatile applications.

Nitrilases represent a distinct class of enzymes that play a pivotal role in catalyzing the hydrolysis of nitrile compounds, leading to the formation of corresponding carboxylic acids. These enzymatic entities have garnered significant attention across a spectrum of industries, encompassing pharmaceuticals, agrochemicals, and fine chemicals. Moreover, their significance has been accentuated by mounting environmental pressures, propelling them into the forefront of biodegradation and bioremediation endeavors.

Not exclusively the activity, but the sweet spot: a dehydrogenase point mutation synergistically boosts activity, substrate tolerance, thermal stability and yield.

Catalytic activity is undoubtedly a key focus in enzyme engineering. The complicated reaction conditions hinder some enzymes from industrialization even though they have relatively promising activity. This has occurred to some dehydrogenases.

Improvement of an enzymatic cascade synthesis of nicotinamide mononucleotide via protein engineering and reaction-process reinforcement.

Enzymatic synthesis of β-nicotinamide mononucleotide (NMN) from D-ribose has garnered widespread attention due to its cheap material, the use of mild reaction conditions, and the ability to produce highly pure products with the desired optical properties. However, the overall NMN yield of this method is impeded by the low activity of rate-limiting enzymes. The ribose-phosphate diphosphokinase (PRS) and nicotinamide phosphoribosyltransferase (NAMPT), that control the rate of the reaction, were engineered to improve the reaction efficacy.

Biosynthesis of Nicotinamide Mononucleotide: Synthesis Method, Enzyme, and Biocatalytic System.

Nicotinamide mononucleotide (NMN) has garnered substantial interest as a functional food product. Industrial NMN production relies on chemical methods, facing challenges in separation, purification, and regulatory complexities, leading to elevated prices. In contrast, NMN biosynthesis through fermentation or enzyme catalysis offers notable benefits like eco-friendliness, recyclability, and efficiency, positioning it as a primary avenue for future NMN synthesis.

Sustainable management and valorization of biomass wastes using synthetic microbial consortia.

In response to the persistent expansion of global resource demands, considerable attention has been directed toward the synthetic microbial consortia (SMC) within the domain of microbial engineering, aiming to address the sustainable management and valorization of biomass wastes. This comprehensive review systematically encapsulates the most recent advancements in research and technological applications concerning the utilization of SMC for biomass waste treatment. The construction strategies of SMC are briefly outlined, and the diverse applications of SMC in biomass wastes treatment are explored, with particular emphasis on its potential advantages in waste degradation, hazardous substances control, and high value-added products conversion.

Identification of a novel thermostable transaminase and its application in L-phosphinothricin biosynthesis.

Transaminase (TA) is a crucial biocatalyst for enantioselective production of the herbicide L-phosphinothricin (L-PPT). The use of enzymatic cascades has been shown to effectively overcome the unfavorable thermodynamic equilibrium of TA-catalyzed transamination reaction, also increasing demand for TA stability. In this work, a novel thermostable transaminase (PtTA) from Pseudomonas thermotolerans was mined and characterized.

Identification of a novel growth-associated promoter for biphasic expression of heterogenous proteins in .

() is one of the most popular cell factories for expressing exogenous proteins and producing useful chemicals. The alcohol oxidase 1 promoter (P) is the most commonly used strong promoter in and has the characteristic of biphasic expression. However, the inducer for P, methanol, has toxicity and poses risks in industrial settings.

Correlation Analysis of Key Residue Sites between Computational-Aided Design Thermostability d-Amino Acid Oxidase and Ancestral Enzymes.

The d-amino acid oxidase (DAAO) from has proven to have great potential for applications due to its excellent catalytic kinetic parameters. However, its poor thermal stability has limited its performance in biocatalysis. Herein, starting from the variant SHVG of DAAO, this study employed a comprehensive computational design approach for protein stability engineering, resulting in positive substitutions at specific sites (A43S, T45M, C234L, E195Y).