Residue profiles of peptides with cholesterol esterase and pancreatic lipase inhibitory activities through virtual screening and sequence analysis.

The detailed characterization of the structural features of peptides targeting cholesterol esterase (CEase) or pancreatic lipase (PPL) will benefit the management of hyperlipidemia and obesity. This study employed the Glide SP (standard precision)-peptide method to predict the binding modes of 202 dipeptides and 203 tripeptides to these targets, correlating residue composition and position with binding energy. Strong preferences for Trp, Phe, and Tyr were observed at all positions of potential inhibitory peptides, whereas negatively charged residues Glu and Asp were disfavored. Notably, Arg and aromatic rings significantly influenced the peptide conformation at the active site. Tripeptide IWR demonstrated the high efficacy, with IC50 values of 0.214 mg/mL for CEase and 0.230 mg/mL for PPL. Five novel IWR scaffold-tetrapeptides exhibited promising inhibitory activity. Non-covalent interactions and energy contributions dominated the formation of stable complexes. Our results provide insights for the development of new sequences or peptide-like molecules with enhanced inhibitory activity.

Shifting Mycobacterial Serine Hydrolase Activity Visualized Using Multi-Layer In-Gel Activity Assays.

The ability of Mycobacterium tuberculosis to derive lipids from the host, store them intracellularly, and then break them down into energy requires a battery of serine hydrolases. Serine hydrolases are a large, diverse enzyme family with functional roles in dormant, active, and reactivating mycobacterial cultures. To rapidly measure substrate-dependent shifts in mycobacterial serine hydrolase activity, we combined a robust mycobacterial growth system of nitrogen limitation and variable carbon availability with nimble in-gel fluorogenic enzyme measurements. Using this methodology, we rapidly analyzed a range of ester substrates, identified multiple hydrolases concurrently, observed functional enzyme shifts, and measured global substrate preferences. Within every growth condition, mycobacterial hydrolases displayed the full, dynamic range of upregulated, downregulated, and constitutively active hydrolases independent of the ester substrate. Increasing the alkyl chain length of the ester substrate also allowed visualization of distinct hydrolase activity likely corresponding with lipases most responsible for lipid breakdown. The most robust expression of hydrolase activity was observed under the highest stress growth conditions, reflecting the induction of multiple metabolic pathways scavenging for energy to survive under this high stress. The unique hydrolases present under these high-stress conditions could represent novel drug targets for combination treatment with current front-line therapeutics. Combining diverse fluorogenic esters with in-gel activity measurements provides a rapid, customizable, and sensitive detection method for mycobacterial serine hydrolase activity.

Implementation of Spore Display in Paenibacillus polymyxa with Different Hydrolytic Enzymes.

Biotechnological processes are essential for producing climate-friendly high-value chemicals or pharmaceutical compounds, which can include steps catalyzed by enzymes. Therefore, establishing new, robust, and cheap enzyme production processes is desirable. One possible way to enhance processes is through the use of the spore display method. Spore display can present heterologous proteins on the surface of bacterial spores, offering numerous advantages in a range of biotechnological applications. This study demonstrates the implementation of the spore display method in Paenibacillus polymyxa, achieved by modifying the spore surface, incorporating an anchoring protein, and attaching green fluorescent protein to it, allowing the visualization of fluorescent spores. Following the initial experiment, a native lipase (Lip3), a heterologous lipase (LipA) from Bacillus subtilis, a native esterase (PnbA) from P. polymyxa, and a lipoyl synthase were expressed during sporulation and displayed on the spore surface. The activity profiles were determined in the temperature range from 4 °C to 70 °C. The PnbA reached its optimum at 4 °C, whereas the LipA from B. subtilis showed 4.4-fold higher activity at 42 °C compared to the control. Furthermore, we explored a possible new technique for the purification of enzymes with the TEV cleavage site between the anchor and the protein of interest. Finally, we showed a not-yet-described side activity of the lipoyl synthase over a wide temperature range.

Neuroprotection by Anethum graveolens (Dill) Seeds and Its Phytocompounds in SH-SY5Y Neuroblastoma Cell Lines and Acellular Assays.

Neurodegeneration diseases (NDs) are a group of complex diseases primarily characterized by progressive loss of neurons affecting mental function and movement. Oxidative stress is one of the factors contributing to the pathogenesis of NDs, including Alzheimer's disease (AD). These reactive species disturb mitochondrial function and accelerate other undesirable conditions including tau phosphorylation, inflammation, and cell death. Therefore, preventing oxidative stress is one of the imperative methods in the treatment of NDs. To accomplish this, we prepared hexane and ethyl acetate extracts of Anethum graveolens (dill) and identified the major phyto-components (apiol, carvone, and dihydrocarvone) by GC-MS. The extracts and major bioactives were assessed for neuroprotective potential and mechanism in hydrogen peroxide-induced oxidative stress in the SH-SY5Y neuroblastoma cell model and other biochemical assays. The dill (extracts and bioactives) provided statistically significant neuroprotection from 0.1 to 30 µg/mL by mitigating ROS levels, restoring mitochondrial membrane potential, reducing lipid peroxidation, and reviving the glutathione ratio. They moderately inhibited acetylcholine esterase (IC50 dill extracts 400-500 µg/mL; carvone 275.7 µg/mL; apiole 388.3 µg/mL), displayed mild anti-Aß1-42 fibrilization (DHC 26.6%) and good anti-oligomerization activity (>40% by dill-EA, carvone, and apiole). Such multifactorial neuroprotective displayed by dill and bioactives would help develop a safe, low-cost, and small-molecule drug for NDs.

Efficient Biodegradation of Multiple Aryloxyphenoxypropionate Herbicides by Corynebacterium sp. Z-1 and the Proposed Degradation Mechanism.

Esterases are crucial for aryloxyphenoxypropionate herbicide (AOPP) biodegradation. However, the underlying molecular mechanisms of AOPP biodegradation by esterases are poorly understood. In the current work, Corynebacterium sp. Z-1 was isolated and found to degrade multiple AOPPs, including quizalofop-p-ethyl (QPE), haloxyfop-p-methyl (HPM), fenoxaprop-p-ethyl (FPE), cyhalofop-butyl (CYB), and clodinafop-propargyl (CFP). A novel esterase, QfeH, which catalyzes the cleavage of ester bonds in AOPPs to form AOPP acids, was identified from strain Z-1. The catalytic activities of QfeH toward AOPPs decreased in the following order: CFP > FPE > CYB > QPE > HPM. Molecular docking, computational analyses, and site-directed mutagenesis indicated the catalytic mechanisms of QfeH-mediated degradation of different AOPPs. Notably, the key residue S159 is essential for the activity of QfeH. Moreover, V222Y, T227M, T227A, A271R, and M275K mutants, exhibiting 2.9-5.0 times greater activity than QfeH, were constructed. This study facilitates the mechanistic understanding of AOPPs bioremediation by esterases.

Recombinant Expression and Characterization of a Novel Thermo-Alkaline Lipase with Increased Solvent Stability from the Antarctic Thermophilic Bacterium Geobacillus sp. ID17.

Lipases are enzymes that hydrolyze long-chain carboxylic esters, and in the presence of organic solvents, they catalyze organic synthesis reactions. However, the use of solvents in these processes often results in enzyme denaturation, leading to a reduction in enzymatic activity. Consequently, there is significant interest in identifying new lipases that are resistant to denaturing conditions, with extremozymes emerging as promising candidates for this purpose. Lip7, a lipase from Geobacillus sp. ID17, a thermophilic microorganism isolated from Deception Island, Antarctica, was recombinantly expressed in E. coli C41 (DE3) in functional soluble form. Its purification was achieved with 96% purity and 23% yield. Enzymatic characterization revealed Lip7 to be a thermo-alkaline enzyme, reaching a maximum rate of 3350 U mg-1 at 50 °C and pH 11.0, using p-nitrophenyl laurate substrate. Notably, its kinetics displayed a sigmoidal behavior, with a higher kinetic efficiency (kcat/Km) for substrates of 12-carbon atom chain. In terms of thermal stability, Lip7 demonstrates stability up to 60 °C at pH 8.0 and up to 50 °C at pH 11.0. Remarkably, it showed high stability in the presence of organic solvents, and under certain conditions even exhibited enzymatic activation, reaching up to 2.5-fold and 1.35-fold after incubation in 50% v/v ethanol and 70% v/v isopropanol, respectively. Lip7 represents one of the first lipases from the bacterial subfamily I.5 and genus Geobacillus with activity and stability at pH 11.0. Its compatibility with organic solvents makes it a compelling candidate for future research in biocatalysis and various biotechnological applications.

Exploring the Esterase Catalytic Activity of Minimalist Heptapeptide Amyloid Fibers.

This paper investigates the esterase activity of minimalist amyloid fibers composed of short seven-residue peptides, IHIHIHI (IH7) and IHIHIQI (IH7Q), with a particular focus on the role of the sixth residue position within the peptide sequence. Through computational simulations and analyses, we explore the molecular mechanisms underlying catalysis in these amyloid-based enzymes. Contrary to initial hypotheses, our study reveals that the twist angle of the fiber, and thus the catalytic site's environment, is not notably affected by the sixth residue. Instead, the sixth residue interacts with the p-nitrophenylacetate (pNPA) substrate, particularly through its -NO2 group, potentially enhancing catalysis. Quantum mechanics/molecular mechanics (QM/MM) simulations of the reaction mechanism suggest that the polarizing effect of glutamine enhances catalytic activity by forming a stabilizing network of hydrogen bonds with pNPA, leading to lower energy barriers and a more exergonic reaction. Our findings provide valuable insights into the intricate interplay between peptide sequence, structural arrangement, and catalytic function in amyloid-based enzymes, offering potentially valuable information for the design and optimization of biomimetic catalysts.

Interaction of plant phenol vanillin with human hemoglobin: A spectroscopic and molecular docking study.

Vanillin is a phenolic aldehyde widely used as a flavouring agent in the food industry. Vanillin has many health benefits and has gained attention in pharmacological industries also, due to its antioxidant properties and non-toxic nature. The interaction of vanillin with human hemoglobin (hHb), an abundant tetrameric heme protein, was investigated by several spectroscopic techniques and molecular modeling methods. UV-visible spectra showed that the binding of vanillin to hHb induces structural changes due to alterations in the micro-environment of hHb. Vanillin quenches the intrinsic fluorescence of hHb by the dynamic mechanism, which was confirmed by both temperature dependent and time resolved fluorescence studies. Vanillin binds spontaneously to hHb at a single site and the binding is stabilized by hydrogen bonds and hydrophobic interactions. The circular dichroism spectra showed that the binding of vanillin altered the secondary structure of hHb due to change in its alpha-helical content. Molecular docking identified the amino acids of hHb involved in binding to vanillin and also that the free energy change of the binding reaction is -5.5 kcal/mol. Thus, our results indicate that vanillin binds spontaneously to hHb at a single site and alters its secondary structure. This will help in understanding the potential use of vanillin and related antioxidants as therapeutic agents in various hematological disorders.

Efficient synthesis and evaluation of therapeutic potential of fluorine containing 2-arylchromen-4-ones.

A large series of 2-arylchromen-4-ones containing from 1 to 3 fluorine atoms or a trifluoromethyl group in the structure was synthesized by condensation of fluorinated 2-hydroxyacetophenones with benzaldehydes in an alkaline medium and subsequent oxidative cyclization of the resulting 2'-hydroxychalcones by action of I2 in DMSO. The cytotoxicity of the obtained compounds was studied in glioblastoma cell line, SNB19, and in a monkey-derived normal kidney epithelium cell line, Vero. In addition, antiglycation activity of the obtained compounds was evaluated. The inhibitory activity of some fluorinated 2-arylchromen-4-ones against acetylcholinesterase, butyrylcholinesterase and carboxylesterase as well their primary antioxidant activity in ABTS and FRAP tests were investigated. Screening of the synthesized compounds for their inhibitory activity against influenza A virus A/Puerto Rico/8/34 (H1N1) in the MDCK cell culture revealed that fluorinated compounds 32, 31 and 39 showed manifest antiviral effects (with IS = 57, 38 and 25 correspondingly) that makes this series of new biologically attractive fluorinated heterocycles promising for further development and in-depth study.

Novel neuroprotective 5,6-dihydropyrido[2',1':2,3]imidazo[4,5-c]quinoline derivatives acting through cholinesterase inhibition and CB2 signaling modulation.

A novel group of 5,6-dihydropyrido [2',1':2,3]imidazo [4,5-c]quinolines was prepared via a microwave assisted one-pot telescopic approach. The synthetic sequence involves the formation of an amine precursor of imidazo [1,2-a]pyridine via condensation and reduction under microwave irradiation. Subsequently, the Pictet-Spengler cyclisation reaction occurs with ketones (cyclic or acyclic) to obtain substituted 5,6-dihydropyrido [2',1':2,3]imidazo [4,5-c]quinolines in excellent yields. The compounds were tested as neuroprotective agents. Observed protection of neuron-like cells, SH-SY5Y differentiated with ATRA, in Parkinson's and Huntington's disease models inspired further mechanistic studies of protective activity against damage induced by 1-methyl-4-phenylpyridinium (MPP+), a compound causing Parkinson's disease. The novel compounds exhibit similar or higher potency than ebselen, an established drug with antioxidant activity, in the cells against MPP + -induced total cellular superoxide production and cell death. However, they exhibit a significantly higher capacity to reduce mitochondrial superoxide and preserve mitochondrial membrane potential. We also observed marked differences between a selected derivative and ebselen in terms of normalizing MPP + -induced phosphorylation of Akt and ERK1/2. The cytoprotective activity was abrogated when signaling through cannabinoid receptor CB2 was blocked. The compounds also inhibit both acetylcholine and butyrylcholine esterases. Overall the data show that novel 5,6-dihydropyrido [2',1':2,3]imidazo [4,5-c]quinoline have a broad cytoprotective activity which is mediated by several mechanisms including mitoprotection.

Exploring the Binding Mechanism of 5-HT7 Specific Benzoxazolone alkyl Piperazinium Derivatives: A Comprehensive Analysis Using Spectroscopic and Computational Approaches.

Recently, the 5-HT7 receptor has achieved greater attention in research fraternity due to the involvement of neurotransmitter serotonin (5-hydroxytryptamine, 5-HT) in several neurological disorders. Targeting this neuroreceptor, we have synthesized six compounds named as butyl-benzoxazolone substituted piperazinium derivatives (BBOP) derivatives, abbreviated as L1-L6. These compounds have been evaluated for their binding interaction with BSA through photophysical and in-silico approaches. The UV absorption of these compounds with BSA at λmax = 280 nm, showed an optical density (O.D.) in the range of 0.5-0.9, i.e., 21%-53% (L1max = 1.4, L5min = 0.7385) at varied concentrations (17 µM-114 µM). For fluorescence studies, the Ksv value varied inversely with temperature, which confirmed the static mechanism of quenching with L1 showing maximum quenching. The parameters (ΔH, ΔS) obtained from the thermodynamic study for interaction between BSA and L1-L6 were correlated with in-silico (molecular docking) data. The in-silico docking study showed hydrophobic and the Van der Waals forces were the most significant forces. Amino acid residues ARG 217 & TRP 213 (Sudlow Site I) and LYS 116 & GLU 125 (Sudlow Site II) of BSA were primarily involved in H-bonding.Furthermore, the catalytic activity of BSA for hydrolyzingdifferent chemical entities have monitored in the presence of L1-L6 through esterase-like assay with p-NPA as a substrate, to get more insight about the interaction with catalytic residues (LYS 414, LYS 413, and TYR 411) in BSA at site II. These findings showed the potential of these 5-HT7 markers as promising ligands with appropriate drug likeliness characteristics.

Structural basis for the inhibition of ßFXIIa by garadacimab.

Activated FXII (FXIIa) is the principal initiator of the plasma contact system and can activate both procoagulant and proinflammatory pathways. Its activity is important in the pathophysiology of hereditary angioedema (HAE). Here, we describe a high-resolution cryoelectron microscopy (cryo-EM) structure of the beta-chain from FXIIa (ßFXIIa) complexed with the Fab fragment of garadacimab. Garadacimab binds to ßFXIIa through an unusually long CDR-H3 that inserts into the S1 pocket in a non-canonical way. This structural mechanism is likely the primary contributor to the inhibition of activated FXIIa proteolytic activity in HAE. Garadacimab Fab-ßFXIIa structure also reveals critical determinants of high-affinity binding of garadacimab to activated FXIIa. Structural analysis with other bona fide FXIIa inhibitors, such as benzamidine and C1-INH, reveals a surprisingly similar mechanism of ßFXIIa inhibition by garadacimab. In summary, the garadacimab Fab-ßFXIIa structure provides crucial insights into its mechanism of action and delineates primary and auxiliary paratopes/epitopes.

Cholinesterase Inhibitory Activity and Molecular Docking Studies of Isocryptolepine-Triazole Adducts.

Due to the rising prevalence of Alzheimer's disease (AD), there is a pressing need for more effective drugs to treat or manage AD's symptoms. Studies have shown that cholinesterase inhibition can improve cognitive and behavioral symptoms associated with AD, by addressing the cholinergic deficit. Based on the recent development of cholinesterase inhibitors with indoloquinoline and triazole moiety, we rationalized that compounds with an isocryptolepine-triazole scaffold may also have the same biological targets. In this study, eighteen previously synthesized isocryptolepine-triazole compounds were assessed for their ability to inhibit acetylcholinesterase (AChE) and butyrylcholine esterase (BChE). The majority of these compounds demonstrated potent selective AChE inhibition. Furthermore, our molecular docking and molecular dynamic simulation studies reveal that the isocryptolepine and triazole moieties are important for the binding of the compounds with the periphery of the AChE's binding pocket. While reductions in molecular weights and lipophilicities may be necessary to improve their pharmacokinetic properties, this work provides valuable insights for designing future AChE inhibitors, based on the novel isocryptolepine-triazole scaffold.

Genome-wide identification of the GDSL-type esterase/lipase protein (GELP) gene family in Ricinus communis and its transcriptional regulation during germination and seedling establishment under different abiotic stresses.

GDSL-type esterase/lipase protein (GELP) genes are crucial in the specialized lipid metabolism, in the responses to abiotic stresses, and in the regulation of plant homeostasis. R. communis is an important oilseed crop species that can sustain growth and productivity when exposed to harsh environmental conditions. Herein, we raised the question of whether the GELP gene family could be involved in the acquisition of R. communis tolerance to abiotic stresses during seed germination and seedling establishment. Thus, we used bioinformatics and transcriptomics to characterize the R. communis GELP gene family. R. communis genome possesses 96 GELP genes that were characterized by extensive bioinformatics, including phylogenetic analysis, subcellular localization, exon-intron distribution, the analysis of regulatory cis-elements, tandem duplication, and physicochemical properties. Transcriptomics indicated that numerous RcGELP genes are readily responsive to high-temperature and salt stresses and might be potential candidates for genome editing techniques to develop abiotic stress-tolerant crops.

Dual Application of p-Nitrophenol Alkanoate-Based Assay for Soil Selection and Screening of Microbial Strains for Bioplastic Degradation.

With an increase in the commercialization of bioplastics, the importance of screening for plastic-degrading strains and microbes has emerged. Conventional methods for screening such strains are time-consuming and labor-intensive. Therefore, we suggest a method for quickly and effectively screening plastic-degrading microbial strains through dual esterase assays for soil and isolated strains, using p-nitrophenyl alkanoates as substrates. To select microbe-abundant soil, the total amount of phospholipid fatty acids (PLFAs) included in each soil sample was analyzed, and esterase assays were performed for each soil sample to compare the esterase activity of each soil. In addition, by analyzing the correlation coefficients and sensitivity between the amount of PLFAs and the degree of esterase activity according to the substrate, it was confirmed that substrate pNP-C2 is the most useful index for soil containing several microbes having esterase activity. In addition, esterase assays of the isolated strains allowed us to select the most active strain as the degrading strain, and 16S rRNA results confirmed that it was Bacillus sp. N04 showed the highest degradation activity for polybutylene succinate (PBS) as measured in liquid culture for 7 days, with a degradation yield of 99%. Furthermore, Bacillus sp. N04 showed degradation activity against various bioplastics. We propose the dual application of p-nitrophenyl alkanoates as an efficient method to first select the appropriate soil and then to screen for plastic-degrading strains in it, and conclude that pNP-C2 in particular, is a useful indicator.

Selective recovery of esterase from Trichoderma harzianum through adsorption: Insights on enzymatic catalysis, adsorption isotherms and kinetics.

In recent years, numerous attempts have been made to develop a low-cost adsorbent for selectively recovering industrially important products from fermentation broth or complex mixtures. The current study is a novel attempt to selectively adsorb esterase from Trichoderma harzianum using cheap adsorbents like bentonite (BT), activated charcoal (AC), silicon dioxide (SiO2), and titanium dioxide (TiO2). AC had the highest esterase adsorption of 97.58% due to its larger surface area of 594.45 m3/g. SiO2 was found to have the highest selectivity over esterase, with an estimated purification fold of 7.2. Interestingly, the purification fold of 5.5 was found in the BT-extracted fermentation broth. The functional (FT-IR) and morphological analysis (SEM-EDX) were used to characterize the adsorption of esterase. Esterase adsorption on AC, SiO2, and TiO2 was well fitted by Freundlich isotherm, demonstrating multilayer adsorption of esterase. A pseudo-second-order kinetic model was developed for esterase adsorption in various adsorbents. Thermodynamic analysis revealed that adsorption is an endothermic process. AC has the lowest Gibbs free energy of -10.96 kJ/mol, which supports the spontaneous maximum adsorption of both esterase and protein. In the desorption study, the maximum recovery of esterase from TiO2 using sodium chloride was 41.34 %. Unlike other adsorbents, the AC-adsorbed esterase maintained its catalytic activity and stability, implying that it could be used as an immobilization system for commercial applications. According to the kinetic analysis, the overall rate of the reaction was controlled by reaction kinetics rather than external mass transfer resistance, as indicated by the Damkohler number.

Tuning of Protein Expression by Promoter Engineering.

The promoter is an essential component of an expression system since it regulates the transcriptional beginning of related genes. The optimal expression level can be achieved by employing a promoter engineering approach. Typically, creating a library of T7 promoters allows for titratable protein expression. In the process of making ß-amino acid (sitagliptin intermediate) from ß-keto ester, esterase from Pseudomonas stutzeri (Est PS) is used to convert the ß-keto ester to ß-keto acid. Subsequently, transaminase from Ilumatobacter coccineus (TAIC) transforms the ß-keto acid to its corresponding ß-amino acid. Here, we describe the optimization of the expression levels of Est PS for the maximum production of sitagliptin intermediate. The different promoter strengths for Est PS were built into the T7 promoters of the pET15b vector. With the help of these new co-expressing entire cells, the expressed enzyme ratio for each enzyme was determined. As the strength of the promoter of Est PS decreases, the expression level also decreases (from 100% to 10%). Conversely, the TAIC expression level is increased. This developed system produced a higher sitagliptin intermediate than enzymes' unoptimized expression level.

Target Enzymes of Origanum majorana and Rosmarinus officinalis Essential Oils in Black Cutworm (Agrotis ipsilon): In Vitro and In Silico Studies.

In this study, in vitro and in silico approaches were employed to assess the toxicity of marjoram (Origanum majorana) and rosemary (Rosmarinus officinalis) essential oils (EOs) to A. ipsilon larvae. The study determined the activities of ATPases in the larvae after treatment with the LC20 and LC70 of each EO. α-esterase and glutathione-S-transferase (GST) activities were also determined after treatment with LC10 and LC30 of each EO. Furthermore, molecular docking was employed to determine the binding affinity of terpinene-4-ol and α-pinene, the major constituents of O. majorana, and R. officinalis EOs, respectively, compared to the co-crystallized ligand of α-esterase, diethyl hydrogen phosphate (DPF). Toxicity assays revealed that O. majorana EO was more toxic than R. officinalis EO to the A. ipsilon larvae at 96 h post-treatment. However, the LC20 and LC70 of the latter significantly inhibited the activity of the Na+-K+ pump at almost all intervals. The same concentrations significantly inhibited the Mg2+/Ca2+-ATPase and Ca2+ pump at 96 h post-treatment. In contrast, O. majorana EO showed a variable effect on the Na+-K+ pump across different time intervals. On the other hand, LC10 and LC30 of both EOs showed varied effects on α-esterase and GST over time. Molecular docking revealed energy scores of -4.51 and -4.29 kcal/mol for terpinene-4-ol and α-pinene, respectively, compared to a score of -4.67 for PDF. Our study demonstrated the toxicity of the tested EOs to A. ipsilon, suggesting their potential efficacy as insecticides.

Discovery, characterization and mechanism of a Microbacterium esterase for key d-biotin chiral intermediate synthesis.

Esterases are crucial biocatalysts in chiral compound synthesis. Herein, a novel esterase EstSIT01 belonging to family V was identified from Microbacterium chocolatum SIT101 through genome mining and phylogenetic analysis. EstSIT01 demonstrated remarkable efficiency in asymmetrically hydrolyzing meso-dimethyl ester [Dimethyl cis-1,3-Dibenzyl-2-imidazolidine-4,5-dicarboxyate], producing over 99% yield and 99% enantiomeric excess (e.e.) for (4S, 5R)-monomethyl ester, a crucial chiral intermediate during the synthesis of d-biotin. Notably, the recombinant E. coli expressing EstSIT01 exhibited over 40-fold higher activity than that of the wild strain. EstSIT01 displays a preference for short-chain p-NP esters. The optimal temperature and pH were 45 °C and 10.0, with Km and kcat values of 0.147 mmol/L and 5.808 s- 1, respectively. Molecular docking and MD simulations suggest that the high stereoselectivity for meso-diester may attribute to the narrow entrance tunnel and unique binding pocket structure. Collectively, EstSIT01 holds great potential for preparing chiral carboxylic acids and esters.

Novel benzene sulfonamide-piperazine hybrid compounds: design, synthesis, antioxidant, enzyme inhibition activities and docking, ADME profiling studies.

Benzene sulfonamides are an important biological substituent for several activities. In this study, hybridization of benzene sulfonamide with piperazine derivatives were investigated for their antioxidant capacity and enzyme inhibitory potencies. Six molecules were synthesized and characterized. DPPH, ABTS, FRAP, CUPRAC, chelating and phosphomolybdemum assays were applied to evaluate antioxidant capacities. Results show that compounds have high antioxidant capacity and compound 4 has the best antioxidant activity among them. Compound 4 has higher antioxidant activity than references for FRAP (IC50: 0.08 mM), CUPRAC (IC50: 0.21 mM) and phosphomolybdenum (IC50: 0.22 mM) assays. Besides this, compound 4 has moderate DPPH and ABTS antioxidant capacity. Furthermore, enzyme inhibition activities of these molecules were investigated against AChE, BChE, tyrosinase, α-amylase and α-glucosidase enzymes. It was revealed that all compounds have good enzyme inhibitory potential except for α-amylase enzyme. The best inhibitory activities were observed for AChE with compound 5 the same value (IC50: 1.003 mM), for BChE with compounds 2 and 5 the same value (IC50: 1.008 mM), for tyrosinase compound 4 (IC50: 1.19 mM), and for α-glucosidase with compound 3 (IC50: 1.000 mM). Docking studies have been conducted with these molecules, and the results correlate well with the inhibitory assays.