Enzymatic Transformation Research Articles

Biotransformation of enrofloxacin-copper combined pollutant in aqueous environments by fungus Cladosporium cladosporioides (CGMCC 40504).

The combined pollution of antibiotics and heavy metals in aqueous environments increases the risk of aquatic ecosystem disruption and the complication of pollutant management. Here, a fungus Cladosporium cladosporioides 11 (CC11) isolated from aquaculture pond sediments possessed effective capacities to biotransform the combined pollution of enrofloxacin (ENR) and copper ion (Cu). ENR and Cu were considerably abated by CC11, and the presence of Cu (30mg/L) promoted the biotransformation efficiency of ENR. The biotranformation of ENR in ENR-Cu co-contamination was associated to ligninolytic enzyme action. The expression of ligninolytic enzymes was enhanced by ENR and ENR-Cu combined pollution. And the increased activities of ligninolytic enzymes confirmed the significant role of enzymatic transformation. Cu played an important role in increasing the expression and activities of ligninolytic enzymes. The expressions of many genes associated with transporters, phosphate assimilation, oxidative phosphorylation, hyperosmotic stress and pectin metabolism were significantly up-regulated when facing Cu-stress, indicating their important roles in determining Cu removal and enhancing Cu-resistance. Additionally, CC11 significantly biotransformed other antibiotic and heavy metal combined pollution. All these results contributed to the applications of CC11 in aqueous environments.

Raman spectroscopic and TEM monitoring of selenite and selenate reduction by the bacterium Azospirillum thiophilum with the formation of selenium(0) nanoparticles: Effects of sulfate

Microbial reduction of selenium oxyanions, highly soluble, mobile and toxic inorganic selenium compounds, to insoluble selenium nanoparticles (Se NPs) is a widely spread phenomenon which is of geochemical, environmental and biotechnological importance. While selenite bioreduction is known for a wide variety of microorganisms, selenate bioreduction is not so common and has mostly been documented for anaerobes, with merely a few reported cases related to aerobic or microaerobic conditions. In some biogenic Se NPs of microbial origin, the presence of sulfur was detected together with selenium in Se NPs, particularly when increased concentrations of sulfate were present in the medium. In this work, the bacterial strain Azospirillum thiophilum BV-S, isolated earlier from a sulfur-containing aqueous environment (Lavrinenko et al. (2010), https://doi.org/10.1099/ijs.0.018853-0), has been shown to reduce selenite to Se NPs also in the presence of 7 mM sulfate in aerobic conditions. Raman spectroscopy was used to monitor the crystallinity and composition of Se NPs formed within the bacterial biomass in the presence of selenite and in the resulting isolated Se NPs, and their spherical morphology was visualised using transmission electron microscopy (TEM). While Se NPs both within the biomass and after isolation gave a typical strong broadened band at 248 cm−1 related to the stretching Se–Se vibrations in amorphous Se0, Raman spectrum of the biomass grown with 1 mM selenite + 7 mM sulfate showed also a weaker band at 348 cm−1 typical of the stretching Se–S mode. The absence of the latter band in Raman spectra of the isolated Se NPs indicates that the Raman-detected Se–S bonds most probably occur in intermediate substances such as selenodiglutathione (GS–Se–SG), an intermediary product in the Painter-type reaction of selenite reduction, which is known to undergo further enzymatic transformations in bacteria resulting in the formation of Se0.Strain A. thiophilum BV-S has also been found, for the first time for bacteria of the genus Azospirillum, to be capable of reducing selenate (SeVIO42–) under static conditions (similar to microaerobic conditions common to habitats of many Azospirillum species in different environments), but not in aerobic conditions, with the formation of Se NPs. The latter, giving a reddish coloration to the bacterial biomass, were characterised by TEM as round-shaped electron-dense structures over bacterial cell images. Raman spectra of the bacterial biomass after cultivation with 5 mM selenate (with or without added sulfate) showed a similar single band at ca. 248 cm−1 in amorphous Se NPs, although 5 mM sulfate slowed down their formation. However, no signs of sulfur covalently bound to Se were observed in Raman spectra in this case. These results indicate a possible negligible role of the selenodisulfide-involving pathway in selenate reduction by A. thiophilum with a noticeable possible interference of sulfate in the process of selenate uptake (i.e., a common transport route) by A. thiophilum cells.

Reversing the Enantioselectivity of Enzymatic Dynamic Kinetic Asymmetric Transformations in the Synthesis of Fused Lactones

Reversing the Enantioselectivity of Enzymatic Dynamic Kinetic Asymmetric Transformations in the Synthesis of Fused Lactones

Recombinant beta-galactosidase derived from Enterobacter cloacae Zjut HJ2001 for efficient biotransformation of galactooligosaccharides

Galactooligosaccharides (GOS), as a type of prebiotics, have excellent physical and chemical properties, and can be widely used in the pharmaceutical and food fields. Recently, the microbial β-galactosidases have gained widespread attentions in industrial GOS production. However, most β-galactosidases from microorganisms have low transgalactosylation activity, resulting in poor GOS yield of enzymatic transformation from lactose. In this paper, a brand new β-galactosidase derived from Enterobacter cloacae Zjut HJ2001 was screened out from soil, and successfully overexpressed, characterized, and mutated by combinatorial alanine-scanning and site-saturation mutagenesis. Compared to the yield of 51.73 % obtained by wild-type β-galactosidase with lactose concentration of 380 g/L, the obtained mutant β-gal-H542V achieved a higher GOS yield of 67.08 %, which was the highest in the reported literature. These results suggested that the developed mutagenesis strategy could improve the transgalactosylation efficiency, and the mutant β-gal-H542V could be regarded as a prospective biocatalyst for GOS industrial manufacturing.

Chemoenzymatic Oxidation of Labdane and Formal Synthesis of Nimbolide.

In nature, basic terpene skeletons are produced and subsequently undergo enzymatic or nonenzymatic oxidative transformations, leading to diverse structural variations. To date, thousands of natural products featuring a variety of oxidation patterns have been isolated solely from the labdane family. This work describes a strategy for the comprehensive introduction of oxidation states into the labdane core by employing a combination of enzyme library screening, directed evolution, and sequential chemical oxidation processes. Furthermore, we showcase the functional viability of our chemoenzymatic approach by accomplishing a formal synthesis of nimbolide, highlighting its potential for streamlining the synthesis of complex natural products.

Green and Enantioselective Synthesis via Cascade Biotransformations: From Simple Racemic Substrates to High-Value Chiral Chemicals.

Asymmetric synthesis of chiral chemicals in high enantiomeric excess (ee) is pivotal to the pharmaceutical industry, but classic chemistry usually requires multi-step reactions, harsh conditions, and expensive chiral ligands, and sometimes suffers from unsatisfactory enantioselectivity. Enzymatic catalysis is a much greener and more enantioselective alternative, and cascade biotransformations with multi-step reactions can be performed in one pot to avoid costly intermediate isolation and minimise waste generation. One of the most attractive applications of enzymatic cascade transformations is to convert easily available simple racemic substrates into valuable functionalised chiral chemicals in high yields and ee. Here, we review the three general strategies to build up such cascade biotransformations, including enantioconvergent reaction, dynamic kinetic resolution, and destruction-and-reinstallation of chirality. Examples of cascade transformations using racemic substrates such as racemic epoxides, alcohols, hydroxy acids, etc. to produce the chiral amino alcohols, hydroxy acids, amines, and amino acids are given. The product concentration, ee, and yield, scalability, and substrate scope of these enzymatic cascades are critically reviewed. To further improve the efficiency and practical applicability of the cascades, enzyme engineering to enhance catalytic activities of the key enzymes using the latest microfluidics-based ultrahigh-throughput screening and artificial intelligence-guided directed evolution could be a useful approach.

Functional characterization of a novel Chlamydomonas reinhardtii hydrolase involved in biotransformation of chloramphenicol

Microalgae-based biotechnology is one of the most promising alternatives to conventional methods for the removal of antibiotic contaminants from diverse water matrices. However, current knowledge regarding the biochemical mechanisms and catabolic enzymes involved in microalgal biodegradation of antibiotics is scant, which limits the development of enhancement strategies to increase their engineering feasibility. In this study, we investigated the removal dynamics of amphenicols (chloramphenicol, thiamphenicol, and florfenicol), which are widely used in aquaculture, by Chlamydomonas reinhardtii under different growth modes (autotrophy, heterotrophy, and mixotrophy). We found C. reinhardtii removed >92 % chloramphenicol (CLP) in mixotrophic conditions. Intriguingly, gamma-glutamyl hydrolase (GGH) in C. reinhardtii was most significantly upregulated according to the comparative proteomics, and we demonstrated that GGH can directly bind to CLP at the Pro77 site to induce acetylation of the hydroxyl group at C3 position, which generated CLP 3-acetate. This identified role of microalgal GGH is mechanistically distinct from that of animal counterparts. Our results provide a valuable enzyme toolbox for biocatalysis and reveal a new enzymatic function of microalgal GGH. As proof of concept, we also analyzed the occurrence of these three amphenicols and their degradation intermediate worldwide, which showed a frequent distribution of the investigated chemicals at a global scale. This study describes a novel catalytic enzyme to improve the engineering feasibility of microalgae-based biotechnologies. It also raises issues regarding the different microalgal enzymatic transformations of emerging contaminants because these enzymes might function differently from their counterparts in animals.

Activity-based metaproteomics driven discovery and enzymological characterization of potential α-galactosidases in the mouse gut microbiome

The gut microbiota offers an extensive resource of enzymes, but many remain uncharacterized. To distinguish the activities of similar annotated proteins and mine the potentially applicable ones in the microbiome, we applied an effective Activity-Based Metaproteomics (ABMP) strategy using a specific activity-based probe (ABP) to screen the entire gut microbiome for directly discovering active enzymes and their potential applications, not for exploring host-microbiome interactions. By using an activity-based cyclophellitol aziridine probe specific to α-galactosidases (AGAL), we successfully identified and characterized several gut microbiota enzymes possessing AGAL activities. Cryo-electron microscopy analysis of a newly characterized enzyme (AGLA5) revealed the covalent binding conformations between the AGAL5 active site and the cyclophellitol aziridine ABP, which could provide insights into the enzyme’s catalytic mechanism. The four newly characterized AGALs have diverse potential activities, including raffinose family oligosaccharides (RFOs) hydrolysis and enzymatic blood group transformation. Collectively, we present a ABMP platform that facilitates gut microbiota AGALs discovery, biochemical activity annotations and potential industrial or biopharmaceutical applications.

Enhancing Efficiency and Selectivity of Electrochemical Regeneration of NADH By Dynamic Operation

The demand for the reduced nicotinamide adenine dinucleotide (1,4-NADH) and reduced nicotinamide adenine dinucleotide phosphate 1,4-NAD(P)H in industrial processes, particularly in the pharmaceutical sector, necessitates their continuous regeneration to mitigate cost implications [1]. In this respect, direct electrochemical cofactor regeneration is a promising alternative, but challenges in achieving optimal selectivity have hindered its widespread industrial integration.This study focuses on advancing the efficiency and selectivity of electrochemical NADH regeneration through dynamic potential inputs, inspired by recent findings demonstrating improved selectivity using dynamic potential inputs instead of steady-state conditions [2]. Various dynamic, pulsed potential profiles are tested and compared with steady-state conditions, revealing distinct advantages in selectivity. Nicotinamide adenine dinucleotide reduction reaction is then coupled with the enzymatic conversion of a substrate (cyclohexenone) to a product (cyclohexanone). In this electroenzymatic biotransformation, 1,4-NADH formed electrochemically is utilized as a second substrate of enzyme enoate reductase. The success of this coupling is evident in the observed formation of the desired product. This research not only contributes to the fundamental understanding of electrochemical co-factor regeneration but also demonstrates its practical application. By combining the advantages of electrochemical and enzymatic reactions, we pave the way for more sustainable and economically viable industrial processes. The presented results mark a significant step towards harnessing the full potential of electrochemical regeneration in facilitating enzymatic transformations with enhanced efficiency and selectivity through a pulsed dynamic operation.

Protein analysis by desorption electrospray ionization mass spectrometry.

This review presents progress made in the ambient analysis of proteins, in particular by desorption electrospray ionization-mass spectrometry (DESI-MS). Related ambient ionization techniques are discussed in comparison to DESI-MS only to illustrate the larger context of protein analysis by ambient ionization mass spectrometry. The review describes early and current approaches for the analysis of undigested proteins, native proteins, tryptic digests, and indirect protein determination through reporter molecules. Applications to mass spectrometry imaging for protein spatial distributions, the identification of posttranslational modifications, determination of binding stoichiometries, and enzymatic transformations are discussed. The analytical capabilities of other ambient ionization techniques such as LESA and nano-DESI currently exceed those of DESI-MS for in situ surface sampling of intact proteins from tissues. This review shows, however, that despite its many limitations, DESI-MS is making valuable contributions to protein analysis. The challenges in sensitivity, spatial resolution, and mass range are surmountable obstacles and further development and improvements to DESI-MS is justified.

Chemo-enzymatic total synthesis: current approaches toward the integration of chemical and enzymatic transformations.

A steadily increasing number of reports have been published on chemo-enzymatic synthesis methods that integrate biosynthetic enzymatic transformations with chemical conversions. This review focuses on the total synthesis of natural products and classifies the enzymatic reactions into three categories. The total synthesis of five natural products: cotylenol, trichodimerol, chalcomoracin, tylactone, and saframycin A, as well as their analogs, is outlined with an emphasis on comparing these chemo-enzymatic syntheses with the corresponding natural biosynthetic pathways.

Regiochemical Analysis of the ProTide Activation Mechanism.

ProTides are nucleotide analogues used for the treatment of specific viral infections. These compounds consist of a masked nucleotide that undergoes in vivo enzymatic and spontaneous chemical transformations to generate a free mononucleotide that is ultimately transformed to the pharmaceutically active triphosphorylated drug. The three FDA approved ProTides are composed of a phosphoramidate (P-N) core coupled with a nucleoside analogue, phenol, and an l-alanyl carboxylate ester. The previously proposed mechanism of activation postulates the existence of an unstable 5-membered mixed anhydride cyclic intermediate formed from the direct attack of the carboxylate group of the l-alanyl moiety with expulsion of phenol. The mixed anhydride cyclic intermediate is further postulated to undergo spontaneous hydrolysis to form a linear l-alanyl phosphoramidate product. In the proposed mechanism of activation, the 5-membered mixed anhydride intermediate has been detected previously using mass spectrometry, but the specific site of nucleophilic attack by water (P-O versus C-O) has not been determined. To further interrogate the mechanism for hydrolysis of the putative 5-membered cyclic intermediate formed during ProTide activation, the reaction was conducted in 18O-labeled water using a ProTide analogue that could be activated by carboxypeptidase Y. Mass spectrometry and 31P NMR spectroscopy were used to demonstrate that the hydrolysis of the mixed anhydride 5-membered intermediate occurs with exclusive attack at the phosphorus center.

Pruned tea biomass plays a significant role in functional food production: A review on characterization and comprehensive utilization of abandon-plucked fresh tea leaves.

Tea is the second largest nonalcoholic beverage in the world due to its characteristic flavor and well-known functional properties in vitro and in vivo. Global tea production reaches 6.397 million tons in 2022 and continues to rise. Fresh tea leaves are mainly harvested in spring, whereas thousands of tons are discarded in summer and autumn. Herein, pruned tea biomass refers to abandon-plucked leaves being pruned in the non-plucking period, especially in summer and autumn. At present, no relevant concluding remarks have been made on this undervalued biomass. This review summarizes the seasonal differences of intrinsic metabolites and pays special attention to the most critical bioactive and flavor compounds, including polyphenols, theanine, and caffeine. Additionally, meaningful and profound methods to transform abandon-plucked fresh tea leaves into high-value products are reviewed. In summer and autumn, tea plants accumulate much more phenols than in spring, especially epigallocatechin gallate (galloyl catechin), anthocyanins (catechin derivatives), and proanthocyanidins (polymerized catechins). Vigorous carbon metabolism induced by high light intensity and temperature in summer and autumn also accumulates carbohydrates, such as soluble sugars and cellulose. The characteristics of abandon-plucked tea leaves make them not ideal raw materials for tea, but suitable for novel tea products like beverages and food ingredients using traditional or hybrid technologies such as enzymatic transformation, microbial fermentation, formula screening, and extraction, with the abundant polyphenols in summer and autumn tea serving as prominent flavor and bioactive contributors.

Regulatory mechanisms of biochar alleviating ammonia inhibition to methanogenesis during long-term operation of anaerobic membrane bioreactor treating swine wastewater

Anaerobic Membrane Bioreactor (AnMBR) is regarded as an advanced technology for the anaerobic digestion (AD) of swine wastewater. However, the high ammonia level in swine wastewater is a crucial issue to inhibit methane production. This study aimed to establish a biochar-assisted AnMBR (BC) to mitigate ammonia inhibition during AD of swine wastewater. As the gradual ammonia loading rate (ALR) increased from 3.0 g/L to 6.0 g/L, BC maintained methane yields ranging from 74.9 % to 48.1 %, which were 9.1 % to 33.3 % higher than those of a regular AnMBR (CT). Moreover, the COD removal efficiency steadily surpassed 80 %. Although the increase in ALR had negative effects on both BC and CT, the presence of biochar enhanced the methanogenic activity of bulk sludge, specifically accelerating processes involved in syntrophic acetate oxidation and CO2-reducing methanogenesis under high ALR conditions. Integrated characterizations using PARAFAC-based EPS analysis, INT-ETS, and WO3 probe-based extracellular electron transfer (EET) activities indicated that biochar stimulated the secretion of humic-like and aromatic protein-like compounds in the TB-EPS, which likely established a redox-active matrix with biochar between VFA oxidation bacteria and CO2-reducing methanogens to facilitate EET. This potentially made the syntrophic VFA oxidation with favorably thermodynamic kinetics under high ammonia stress. Furthermore, the analysis of microbial community coupling prediction of functional genes associated with methanogenic pathways revealed that as ALR increased, the presence of biochar re-shaped the microbial communities by enriching electro-active Syntrophomonas and Methanosarcina/Methanospirillum, which mainly triggering EET-based syntrophic methanogenesis by accelerating the intracellular enzymatic transformation from Formylmethanofuran to 5,10-Methylenetetrahydromethanopterin process. We expect the major findings of this study provide new insights to understand the mechanisms of biochar alleviating ammonia inhibition during long-term operation of AnMBR.

Enzyme-Driven LC-HRMS Approach for Specific Recognition of 12α-Hydroxy Bile Acids.

The synthesis of 12α-hydroxylated bile acids (12HBAs) and non-12α-hydroxylated bile acids (non-12HBAs) occurs via classical and alternative pathways, respectively. The composition of these BAs is a crucial index for pathophysiologic assessment. However, accurately differentiating 12HBAs and non-12HBAs is highly challenging due to the limited standard substances. Here, we innovatively introduce 12α-hydroxysteroid dehydrogenase (12α-HSDH) as an enzymatic probe synthesized by heterologous expression in Escherichia coli, which can specifically and efficiently convert 12HBAs in vitro under mild conditions. Coupled to the conversion rate determined by liquid chromatography-high resolution mass spectrometry (LC-HRMS), this enzymatic probe allows for the straightforward distinguishing of 210 12HBAs and 312 non-12HBAs from complex biological matrices, resulting in a BAs profile with a well-defined hydroxyl feature at the C12 site. Notably, this enzyme-driven LC-HRMS approach can be extended to any molecule with explicit knowledge of enzymatic transformation. We demonstrate the practicality of this BAs profile in terms of both revealing cross-species BAs heterogeneity and monitoring the alterations of 12HBAs and non-12HBAs under asthma disease. We envisage that this work will provide a novel pattern to recognize the shift of BA metabolism from classical to alternative synthesis pathways in different pathophysiological states, thereby offering valuable insights into the management of related diseases.

Cordycepin Triphosphate as a Potential Modulator of Cellular Plasticity in Cancer via cAMP-Dependent Pathways: An In Silico Approach.

Cordycepin, or 3'-deoxyadenosine, is an adenosine analog with a broad spectrum of biological activity. The key structural difference between cordycepin and adenosine lies in the absence of a hydroxyl group at the 3' position of the ribose ring. Upon administration, cordycepin can undergo an enzymatic transformation in specific tissues, forming cordycepin triphosphate. In this study, we conducted a comprehensive analysis of the structural features of cordycepin and its derivatives, contrasting them with endogenous purine-based metabolites using chemoinformatics and bioinformatics tools in addition to molecular dynamics simulations. We tested the hypothesis that cordycepin triphosphate could bind to the active site of the adenylate cyclase enzyme. The outcomes of our molecular dynamics simulations revealed scores that are comparable to, and superior to, those of adenosine triphosphate (ATP), the endogenous ligand. This interaction could reduce the production of cyclic adenosine monophosphate (cAMP) by acting as a pseudo-ATP that lacks a hydroxyl group at the 3' position, essential to carry out nucleotide cyclization. We discuss the implications in the context of the plasticity of cancer and other cells within the tumor microenvironment, such as cancer-associated fibroblast, endothelial, and immune cells. This interaction could awaken antitumor immunity by preventing phenotypic changes in the immune cells driven by sustained cAMP signaling. The last could be an unreported molecular mechanism that helps to explain more details about cordycepin's mechanism of action.

Study on the anti–inflammatory effect of 3–(4–hydroxyphenyl) propionic acid in an in vitro LPS–stimulated acute kidney inflammation model

Acute kidney injury (AKI) is a syndrome defined by a rapid decrease in glomerular filtration that can be caused by sepsis, ischemia/reperfusion injury (IRI), or nephrotoxic drugs. Human microbiota makes significant contributions to human health by enzymatic transformation of such active substances and the release of molecules such as 3–4 hydroxyphenyl propionic acid (4–HPPA). Biological effects of 4–HPPA such as anti–inflammatory and antioxidant have been reported in many studies. The aim of the research is to reveal the anti–inflammatory activity of 4–HPPA, one of the microbiota products of flavonoids (especially naringin) found in many fruits, in an in vitro LPS (lipopolysaccharide) stimulated kidney inflammation model. HEK 293 kidney cells of human origin were used as material in the research. The trial consisted of 4 groups: control group, LPS group, 4–HPPA group and 4–HPPA+LPS group. LPS and 4–HPPA were applied to the cells at different concentrations for 24 hours. Effective concentrations of LPS and 4–HPPA were investigated by MTT viability test. Finally, IL–1β, TNF–α and NFkβ gene expression analyzes responsible for inflammatory responses were investigated by qRT–PCR method. According to the findings, after 24 hours of incubation, LPS at 2.5 ng·mL-1 and 4–HPPA at 6.25 μg·mL-1 were determined to be effective concentrations for the experiment. Again, it was observed that 4–HPPA downregulated LPS–induced IL–1β, TNF–α and NFkβ gene expressions by 7, 42 and 40%, respectively. According to the data obtained from the research, it was revealed that 4–HPPA had effective anti–inflammatory properties in the in vitro LPS–stimulated kidney inflammation model. However, it was concluded that in vivo and more advanced molecular methods are needed to fully elucidate the issue.

The β-subunit of tryptophan synthase is a latent tyrosine synthase.

Aromatic amino acids and their derivatives are diverse primary and secondary metabolites with critical roles in protein synthesis, cell structure and integrity, defense and signaling. All de novo aromatic amino acid production relies on a set of ancient and highly conserved chemistries. Here we introduce a new enzymatic transformation for L-tyrosine synthesis by demonstrating that the β-subunit of tryptophan synthase-which natively couples indole and L-serine to form L-tryptophan-can act as a latent 'tyrosine synthase'. A single substitution of a near-universally conserved catalytic residue unlocks activity toward simple phenol analogs and yields exclusive para carbon-carbon bond formation to furnish L-tyrosines. Structural and mechanistic studies show how a new active-site water molecule orients phenols for a nonnative mechanism of alkylation, with additional directed evolution resulting in a net >30,000-fold rate enhancement. This new biocatalyst can be used to efficiently prepare valuable L-tyrosine analogs at gram scales and provides the missing chemistry for a conceptually different pathway to L-tyrosine.

Gas chromatography-mass spectrometry characterization of the aroma of Ossetian cheeses

The aim of the study was the qualitative and quantitative determination of volatile aroma compounds and their formation pathways in brine Ossetian cheeses. Volatile components of cheeses were isolated by steam distillation and extraction with dichloromethane, with their subsequent determination and quantification by gas chromatography-mass spectrometry. The results of the analysis are presented according to the structural classes of the main chemical components and the corresponding microbial metabolic processes. Four processes were found to be the main contributors to flavor formation: lipolysis, proteolysis, glycolysis, and a number of oxidative enzymatic transformations. Lipolysis of the fatty fraction of cheeses is a source of formation of volatile carboxylic acids and their esters. Proteolysis of the casein fraction yields branched alcohols, aldehydes, and a number of aromatic and heteroaromatic compounds. Glycolysis of the carbohydrate fraction is a source of ethanol formation, which is the main cause of the dominance of ethyl esters in the ester fraction. Redox enzymatic transformations mainly determine the biosynthesis of unbranched aldehydes, ketones and lactones. A clear distinction between retail and homemade cheeses was observed, due to the different technological approaches to the cheese preparation. The structuralchemical and quantitative evolution of the volatile composition of the studied cheese samples during ripening is tentatively shown. From the authors’ point of view, the aromatic composition of the Tib cheese sort is the most consistent with the Ossetian cheese standard. This study represents the first gas chromatographic study of Ossetian cheeses and aims to create objective criteria for controlling technological processes and product quality during production and storage in the food industry.

Chemoenzymatic synthesis of macrocyclic peptides and polyketides via thioesterase-catalyzed macrocyclization.

Chemoenzymatic strategies that combine synthetic and enzymatic transformations offer efficient approaches to yield target molecules, which have been increasingly employed in the synthesis of bioactive natural products. In the biosynthesis of macrocyclic nonribosomal peptides, polyketides, and their hybrids, thioesterase (TE) domains play a significant role in late-stage macrocyclization. These domains can accept mimics of native substrates in vitro and exhibit potential for use in total synthesis. This review summarizes the recent advances of TE domains in the chemoenzymatic synthesis for these natural products that aim to address the common issues in classical synthetic approaches and increase synthetic efficiencies, which have the potential to facilitate further pharmaceutical research.