A5055715828- Enzyme Catalysis and Immobilization
- Microbial Metabolic Engineering and Bioproduction
- Chemical Synthesis and Analysis
- Innovative Microfluidic and Catalytic Techniques Innovation
- Silk-based biomaterials and applications
- Catalysis for Biomass Conversion
- Synthetic Organic Chemistry Methods
- Amino Acid Enzymes and Metabolism
- Nanofabrication and Lithography Techniques
- Peptidase Inhibition and Analysis
- Click Chemistry and Applications
- Biochemical and Molecular Research
- Neurobiology and Insect Physiology Research
- Biochemical and Structural Characterization
- Microbial Natural Products and Biosynthesis
- Enzyme Structure and Function
- Carbohydrate Chemistry and Synthesis
- Adenosine and Purinergic Signaling
- Pharmacogenetics and Drug Metabolism
- Steroid Chemistry and Biochemistry
- Biofuel production and bioconversion
- Cancer-related gene regulation
- Viral Infectious Diseases and Gene Expression in Insects
- Cancer, Hypoxia, and Metabolism
- Advanced Cellulose Research Studies
University of Manchester
2019-2024
Czech Academy of Sciences, Institute of Biotechnology
2021
University of Exeter
2016-2021
The state-of-the-art of biocatalytic amide bond formation is discussed with the help a manually curated database enzymatic amidation reactions.
Abstract Carboxylic acid reductase enzymes (CARs) meet the demand in synthetic chemistry for a green and regiospecific route to aldehydes from their respective carboxylic acids. However, relatively few of these have been characterized. A sequence alignment with members ANL (Acyl‐CoA synthetase/ NRPS adenylation domain/Luciferase) superfamily shed light on CAR functional dynamics. Four unstudied were selected by using phylogenetic analysis known hypothetical CARs, first time, thorough...
Abstract Carboxylic acid reductases (CARs) are biocatalysts of industrial importance. Their properties, especially their poor stability, render them sub-optimal for use in a bioindustrial pipeline. Here, we employed ancestral sequence reconstruction (ASR) – burgeoning engineering tool that can identify stabilizing but enzymatically neutral mutations throughout protein. We used three-algorithm approach to reconstruct functional ancestors the Mycobacterial and Nocardial CAR1 orthologues....
Monoacylated diamines are common motifs present in pharmaceuticals, agrochemicals, and natural products. However, the synthesis of these compounds often requires selective protection/deprotection steps leading to waste poor atom economy. Here we describe truncation a carboxylic acid reductase (CAR) yielding an adenylation domain capable amide bond formation under aqueous conditions. This truncated construct had higher amidation activity compared full-length enzyme. The reaction is mediated...
N-alkanoyl-N-methylglucamides (MEGAs) are non-toxic surfactants widely used as commercial ingredients, but more sustainable syntheses towards these compounds highly desirable. Here, we present a biocatalytic route MEGAs and analogues using truncated carboxylic acid reductase construct tailored for amide bond formation (CARmm-A). CARmm-A is capable of selective without the competing esterification reaction observed in lipase catalysed reactions. A kinase was implemented to regenerate ATP from...
Despite the increasing use of biocatalysis for organic synthesis, there are currently no databases that adequately capture synthetic biotransformations. The lack a database prevents accelerating biocatalyst characterization efforts from being leveraged to quickly identify candidate enzymes reactions or cascades, slowing their development. RetroBioCat Database (available at retrobiocat.com) addresses this gap by capturing information on biotransformations and providing an analysis platform...
Multi-step enzyme reactions offer considerable cost and productivity benefits. Process models a route to understanding the complexity of these reactions, allow for their optimization. Despite increasing prevalence multi-step biotransformations, there are few examples process reactions. From toolbox characterized parts, we demonstrate construction model seven enzyme, three step biotransformation using isolated enzymes. Enzymes cofactor regeneration were employed make this in vitro reaction...
Spider silk spidroins consist of long repetitive protein strands, flanked by globular terminal domains. The domains are often omitted in recombinant spidroins, but thought to be essential for the spiders' natural spinning process. Mimicking this process could an step towards producing strong synthetic spider silk. Here we describe production a range mini-spidroins with both domains, and characterize their response number biomimetic triggers. Our results suggest that which able form micelles...
Protein-based adhesives could have several advantages over petroleum-derived alternatives, including substantially lower toxicity, smaller environmental footprint, and renewable sourcing. Here, we report that non-covalently crosslinked bovine serum albumin recombinant spider silk proteins high adhesive strength on glass (8.53 6.28 MPa, respectively) other transparent substrates. Moreover, the visible transparency showed no apparent degradation a period of months. The mechanism adhesion was...
Enzyme-CBM fusion constructs immobilised on wood-derived cellulose scaffolds: a sustainable approach for continuous flow biocatalysis.
Biocatalysis is important in the discovery, development, and manufacture of pharmaceuticals. However, identification enzymes for target transformations interest requires major screening efforts. Here, we report a structure-based computational workflow to prioritize protein sequences by score based on predicted activities substrates, thereby reducing resource-intensive laboratory-based biocatalyst screening. We selected imine reductases (IREDs) as class biocatalysts illustrate application...
We report the development of an engineered aminotransferase for synthesis a key chiral intermediate anti-HIV drug Lenacapavir. Due to sterically demanding nature ketone substrate, substrate walking approach was adopted during directed evolution unlock desired activity starting from parent template (TA25) with no observable target reaction. Introduction 6 mutations into TA25 over 4 rounds led that affords amine product 90% conversion and >99% e.e. in favor S-enantiomer. The enzyme serves...
Nucleosides functionalized at the 2'-position play a crucial role in therapeutics, serving as both small-molecule drugs and modifications therapeutic oligonucleotides. However, synthesis of these molecules often presents substantial synthetic challenges. Here we present an approach to 2'-functionalized nucleosides based on enzymes from purine nucleoside salvage pathway. Initially, active-site variants deoxyribose-5-phosphate aldolase were generated for highly stereoselective...
Synthetic biology has huge potential to produce the next generation of advanced materials by accessing previously unreachable (bio)chemical space. In this prospective review, we take a snapshot current activity in rapidly developing area, focussing on prominent examples for high-performance applications such as those required protective and aerospace sector. The continued growth emerging field will be facilitated convergence expertise from range diverse disciplines, including molecular...
Enzymes are increasingly combined into multienzyme systems for cost and productivity benefits. Further advantages can be gained through the use of immobilized enzymes, allowing continuous biotransformations in flow. However, optimization such is challenging, particularly where enzymes used. Here, we meet this challenge using both mechanistic empirical modeling to optimize a reaction involving reductive aminase glucose dehydrogenase biocatalytic amination Crucially, construction model was...
Abstract A new carboxyl esterase, AF-Est2, from the hyperthermophilic archaeon Archaeoglobus fulgidus has been cloned, over-expressed in Escherichia coli and biochemically structurally characterized. The enzyme high activity towards short- to medium-chain p -nitrophenyl carboxylic esters with optimal valerate ester. AF-Est2 good solvent pH stability is very thermostable, showing no loss of after incubation for 30 min at 80 °C. 1.4 Å resolution crystal structure reveals Coenzyme (CoA) bound...
Nucleosides functionalized at the 2′-position play a crucial role in therapeutics, serving as both small molecule drugs and modifications therapeutic oligonucleotides. However, synthesis of these molecules often presents significant synthetic challenges. In this study, we present an approach to 2′-functionalized nucleosides based on enzymes from purine nucleoside salvage pathway. Initially active-site variants DERA aldolase were generated for highly stereoselective D-ribose-5-phosphate...
Osmolytes protect microbial cells against temperature, osmolarity and other stresses. The osmolyte cyclic 2,3-diphosphoglycerate, originally isolated from the thermophilic archaeon Methanothermus fervidus , naturally protects cellular proteins under extreme conditions. biosynthetic pathway for 2,3-diphosphoglycerate has been introduced into bacterium Thermus thermophilus . two enzymes in this synthetic pathway, 2-phosphoglycerate kinase diphosphoglycerate synthetase, were incorporated a...
Abstract N ‐alkanoyl‐ ‐methylglucamides (MEGAs) are non‐toxic surfactants widely used as commercial ingredients, but more sustainable syntheses towards these compounds highly desirable. Here, we present a biocatalytic route MEGAs and analogues using truncated carboxylic acid reductase construct tailored for amide bond formation (CAR mm ‐A). CAR ‐A is capable of selective without the competing esterification reaction observed in lipase catalysed reactions. A kinase was implemented to...
We report the development of an engineered P450 monooxygenase that mediates a chemo- and stereo-selective alkene epoxidation to generate key chiral precursor anti-tuberculosis drug delamanid. Screening in-house panel led identification BM3 variant, containing five mutations, with activity for target transformation. Over single round laboratory evolution gene shuffling, three further beneficial mutations were introduced leading order magnitude increase in de-sired activity, total turnover...