Active Binding Sites Research Articles

Synthesis, anticancer evaluation and in silico studies of novel N‐substituted arylidenethiazolidine‐2,4‐dione derivatives as adenosine monophosphate‐activated protein kinase activators

AbstractDesign and development of AMP‐activated protein kinase (AMPK) activator emerged as a potential therapeutic approach for various types of cancers. In this context, thiazolidine 2,4‐dione was invariably found as an important skeleton for the development of new lead compounds. The present study described the synthesis and antitumor evaluation of new hybrids of N‐substituted arylidenethiazolidine‐2,4‐diones as AMPK activators. The in vitro results revealed that several of newly prepared compounds exhibited significant anticancer activity against human prostate cancer (PC3) and breast cancer (MDMB‐231) cell growths with IC50 in the range of 2–10 μM. Particularly, molecular hybridization of thiazolidine 2,4‐dione with N‐2‐(4‐(trifluoromethyl)phenyl)ethanol and azaindole (compound 16) was the most effective among the series against both PC3 and MDMB‐231 cell lines with IC50 4.28 and 2.5 μM, respectively. Western blot analysis of these thiazolidine 2,4‐dione hybrids showed increased (p)‐AMPK level in the PC‐3 cells indicating direct activation of AMPK. The docking studies at the interface of activator binding site of the AMPK reinforced the in vitro results of potent compounds 13, 16, and 25 having low docking scores −9.0, −9.5, and −9.1 Kcal/mol, respectively.

Synthesis, biological evaluation and molecular docking study of pyrimidine linked thiazolidinedione derivatives as potential antimicrobial and antitubercular agents

The design and development of novel antimicrobial agents are highly desired to combat the emergence of medication resistance against microorganisms that cause infections. A series of new pyrimidine-linked thiazolidinedione derivatives (5a-j) were synthesized, characterized, and their antimicrobial properties assessed in the current investigation. Here, novel pyrimidine-linked thiazolidinedione compounds were designed using the molecular hybridization approach. Elemental and spectral techniques were used to determine the structures of the synthesized hybrids. The majority of compounds showed encouraging antibacterial properties. Among the active compounds, 5g, 5i, and 5j showed 1.85, 1.15, and 1.38 times the activity of streptomycin against S. aureus, respectively, with MIC values of 6.4, 10.3, and 8.6 µM. With MIC values of 10.8, 21.9, and 15.4 µM, respectively, the compounds 5g, 5i, and 5j showed 2.14, 1.05, and 1.50 times the activity of linezolid against the methicillin-resistant S. aureus (MRSA) strain. Furthermore, when compared to the reference medications, compounds 5g, 5i, and 5j demonstrated broad-range antimicrobial efficacy against all tested strains of bacteria and fungus. Out of all the compounds that were investigated, compounds 5g, 5i, and 5j showed noteworthy anti-tubercular activity. 5g is the most effective, 1.59 times more effective than reference drug isoniazid. To anticipate the binding manner, the synthesized potent compounds were subjected to molecular docking into the active binding site of MRSA and the mycobacterial membrane protein large 3 (MmpL3) protein. The compounds 5g, 5i, and 5j may eventually serve as lead compounds in the search for antimicrobial and anti-TB therapeutic agents.

Paynantheine more effectively reverses multidrug resistance in malignant EPG85.257RDB and MCF7MX cells than morphine and speciociliatine.

The possible interactions of morphine, paynantheine and speciociliatine alkaloids with ATP-binding cassette (ABC) transporters was investigated. The compounds were docked against ABCG2 and ABCB1 to predict the binding mode of alkaloids in active binding sites. The cytotoxicity of morphine, paynantheine and speciociliatine for EPG85.257RDB and MCF7MX cells was determined and ABCB1 and ABCG2 gene and protein expression were determined. The binding score of paynantheine to ABCB1 was higher in the docking studies. Paynantheine and speciociliatine had similar binding scores to ABCB1, but higher binding scores to ABCG2 than did morphine. Paynantheine and speciociliatine were more effective against MCF7MX and EPG85.257RDB cells and showed greater cyctotoxicity in the MTT assay. The effect of morphine and paynantheine on the ABCB1 gene and protein expression suggests these compounds can reduce resistance in cancer patients, but that speciociliatine may not be a suitable candidate because of its increased ABCB1 expression while speciociliatine decreased the expression of ABCG2 in MCF7MX cells. This indicates that speciociliatine is a better candidate for reducing drug resistance in this cell line. Structural modification, drug-metabolizing enzymes and differences in the binding sites could cause functional differences between these compounds.

Ultrasensitive detection of miRNA based on single-walled carbon nanotube@MoS2 field-effect transistor regulated by atomic layer deposition

MicroRNAs (miRNAs) act as diagnostic markers for various diseases, and achieving ultra-sensitive detection of miRNAs has an important effect on early disease diagnosis. Field-effect transistor (FET) biosensor exhibits superior sensitivity compared to conventional clinical detection instruments in detecting biomolecules. However, developing high-sensitive FET biosensors for detecting microRNAs remains challenging. In this study, we developed a FET biosensor using MoS2 and a network film of semiconductor single-walled carbon nanotube (SWCNT) as channel materials. The MoS2 cycles were precisely controlled and optimized by atom layer deposition (ALD), and the optimized 3-cycle device exhibited the best performance. This adjustment is beneficial for enhancing the charge transfer between SWCNT and MoS2, as well as improving the on/off ratio and mobility of FET devices. Additionally, the SWCNT@MoS2 structure offers a larger specific surface area, providing more active binding sites on the sensing interface. These two favorable factors contribute to the realization of linear detection miRNA-21 from 1 × 10−17 to 1 × 10−10 mol L−1 in biosensors. The theoretical detection limit of the biosensor is 1.9 × 10−18 mol L−1. This work develops a novel approach to achieve highly sensitive detection of miRNA for high-performance FET biosensors.

Design, Synthesis, and in-silico Studies of New Disubstituted Benzimidazoles: Evaluation of In-vitro Anticancer and Antimicrobial Activity

A series of thirteen new benzimidazole scaffolds 4(a-m) was synthesized by nucleophilic substitution of 2-chloro- N-(aryl)-acetamides 3(a-m) with 2-(substituted phenyl)-1H-benzo[d]imidazoles 2(a-e) in N.N-Dimethylformamide. All the synthesized compounds were tested on the MCF-7 cell line, followed by antibacterial and antifungal activity. The anticancer activity was determined using MTT assay technique, whereas the antimicrobial activity was carried out using two-fold serial dilution technique. Among all the tested compounds, compound 4b exhibited the highest anticancer and antimicrobial activity, comparable to the standard drug. Finally, the binding mode analysis of the highly active compounds was performed at the active binding sites of the co-crystallized structure of the MCF-7 cell line (PDB ID 1ZXM) and glucosamine-6-phosphate synthase (PDB ID 2VF5). In silico absorption, distribution, metabolism, and excretion studies were also carried out.. KEYWORDS :Benzimidazoles, Cytotoxicity, in silico studies, MCF-7 cell lines, MTT assay.

Polycarboxyl ionic liquid functionalized Yb-MOFs nanoballs based dual-wavelength responsive photoelectrochemical aptasensor for the simultaneous determination of AFB1 and OTA

Developing an accurate and precise approach for the simultaneous detection of ochratoxin A (OTA) and aflatoxin B1 (AFB1) is significant for food safety surveillance. Herein, a photoelectrochemical sensing platform was constructed based on polycarboxylic ionic liquid functionalized metal-organic framework integrated with gold nanoparticles (Yb-MOFs@AuNPs). Sulfhydryl functionalized hairpin DNA (hDNA) was immobilized on a Yb-MOFs@AuNPs modified glassy carbon electrode (GCE) surface through Au-S bond. After blocking residual active binding sites with BSA, gold nanoparticles-labeled AFB1 aptamer (AuNPs-Apt 1) and gold nanorods-labeled OTA aptamer (AuNRs-Apt 2) were introduced to construct a photoelectrochemical aptasensor for the simultaneous determination of AFB1 and OTA. Due to the surface plasmon resonance effect and the nanometer size effect of gold nanomaterials, the photoelectrochemical aptasensor can output photocurrent responses as being excited with different wavelengths at 520 nm and 808 nm, respectively. When the AFB1 and OTA concentration in the range of 0.001–50.0 ng mL−1, a good linear relationship between the photocurrent difference (ΔI) before and after recognizing targets and the logarithm of AFB1 or OTA concentration was obtained. The detection limits for AFB1 and OTA were 0.40 pg mL−1 and 0.19 pg mL−1, respectively. AFB1 and OTA in corn samples were detected simultaneously by the photoelectrochemical aptasensor.

Deciphering the features and functions of serine/arginine protein kinases in bread wheat

Serine/arginine protein kinases (SRPKs) are members of the serine-threonine kinase family that phosphorylate the Serine/arginine-rich (SR) proteins involved in alternate splicing. They are reported in various eukaryotes including mammals, and in a few plants, but seldom explored in important crop species. Herein, we identified a total of nine TaSRPK genes from all three subgenomes (A, B, and D) of a staple crop Triticum aestivum, and phylogenetically classified them into two groups. The TaSRPKs have conserved gene architecture with four exons. Each TaSRPK protein consists of a characteristic protein kinase domain having an active site and ATP binding region. The occurrence of diverse cis-regulatory elements in the promoter region, and interaction with assorted groups of transcription factors and miRNAs exhibited their divergent functions. Differential expression of certain TaSRPKs in vegetative and reproductive tissues and in the presence of fungal pathogens and various abiotic stress conditions further assured their association during development processes and stress response. Our study highlighted the importance of TaSRPKs, which might be useful for their detailed characterization in future research.

Multifunctional Ag2O/chitosan nanocomposites synthesized via sol-gel with enhanced antimicrobial, and antioxidant properties: A novel food packaging material

In this study, a single step in situ sol-gel method was used to syntheses nanocomposite films using chitosan (CS) as the basis material, with the addition of silver oxide nanoparticles (Ag2O) at several weight percentages (5 %, 10 %, and 15 % Ag2O/CS). The structural characteristics of Ag2O/CS films were investigated using a range of analytical techniques. The presence of the primary distinctive peaks of chitosan was verified using FTIR spectra analysis. However, a minor displacement was observed in these peaks due to the chemical interaction occurring with silver oxide molecules. XRD analysis demonstrated a significant increase in the crystallinity of chitosan when it interacted with metal oxide nanoparticles. Furthermore, it is believed that the interaction between silver oxide and the active binding sites of chitosan is responsible for the evenly dispersed clusters shown in the micrographs of the chitosan surface, as well as the random aggregations within the pores. EDS technique successfully identified the presence of distinctive silver signals within the nanocomposite material, indicating the successful absorption of silver into the surface of the polymer. The developed Ag2O/CS nanocomposite showed promising antibacterial activity against Gram-negative (Escherichia coli and Pseudomonas aeruginosa) and Gram-positive bacteria (Bacillus subtilis, Enterococcus faecalis and Staphylococcus aureus). Also, Ag2O/CS nanocomposite exhibited marked antifungal activity against Candida albicans, Aspergillus flavus, A. fumigatus, A. niger, and Penicillium chrysogenum. The antioxidant activity of the developed nanocomposite films was studied by ABTS radical scavenging. The highest antioxidant and antibacterial properties were achieved by including 15 % silver oxide into the chitosan. Therefore, our finding indicate that chitosan‑silver oxide nanocomposites exhibits significant potential as a viable material for application in several sectors of the food packaging industry.

Synthesis of magnetic bifunctional monomer molecularly imprinted polymers for highly selective separation of paclitaxel from Taxus cuspidata

Paclitaxel (PTX), an active anticancer compound derived from Taxus cuspidata, possesses considerable economic value. Given the abundance of structural analogs, the selective separation of PTX from T. cuspidata presents a formidable challenge. In this study, the bifunctional monomers magnetic molecular imprinted polymers (MMIPs) with multiple active sites were synthesized using Fe3O4 as the magnetic carrier, PTX as the template, 4-vinylpyridine (4-VP) and methacrylic acid (MAA) as the first-stage and the second-stage pre-assembly functional monomer, respectively. The results indicated that MMIPs exhibited excellent magnetic recovery capability and high adsorption capacity for PTX. Furthermore, the adsorption behaviors were more consistent with Langmuir isotherm model and pseudo-second-order kinetic model. The MMIPs possessed significantly higher selectivity for PTX than five structural analogs (Docetaxel, Baccatin III, 10-Deacetylbaccatin III, and Glucose) and recyclability for up to six cycles. Additionally, the adsorption mechanism analysis indicated that the excellent performance of MMIPs primarily arises from the multiple active binding sites involving hydrogen bonding and π-π stacking. Finally, MMIPs were used for PTX recognition and separation in T. cuspidata extracts. This study provides an efficient method and valuable theoretical basis for the highly selective separation of PTX from T. cuspidata.

Rapid and Ultrasensitive Colorimetric Biosensors for Onsite Detection of Escherichia coli O157:H7 in Fluids.

This study presents a breakthrough in the field of onsite bacterial detection, offering an innovative, rapid, and ultrasensitive colorimetric biosensor for the detection of Escherichia coli (E. coli) O157:H7, using chemically modified melamine foam (MF). Different from conventional platforms, such as 96-well plates and fiber-based membranes, the modified MF features a macroporous reticulated three-dimensional (3D) framework structure, allowing fast and free movement of large biomolecules and bacteria cells through the MF structure in every direction and ensuring good accessibility of entire active binding sites of the framework structure with the target bacteria, which significantly increased sensitive and volume-responsive detection of whole-cell bacteria. The biosensing platform requires less than 1.5 h to complete the quantitative detection with a sensitivity of 10 cfu/mL, discernible by the naked eye, and an enhanced sensitivity of 5 cfu/mL with the help of a smartphone. Following a short enrichment period of 1 h, the sensitivity was further amplified to 2 cfu/mL. The biosensor material is volume responsive, making the biosensing platform sensitivity increase as the volume of the sample increases, and is highly suitable for testing large-volume fluid samples. This novel material paves the way for the development of volume-flexible biosensing platforms for the record-fast, onsite, selective, and ultrasensitive detection of various pathogenic bacteria in real-world applications.

ПРИМЕНЕНИЕ СОРБЕНТА ДЛЯ ТВЕРДОФАЗНОЙ ЭКСТРАКЦИИ НА ОСНОВЕ ГИПРОМЕЛЛОЗЫ ДЛЯ КОЛИЧЕСТВЕННОГО ОПРЕДЕЛЕНИЯ КИСЛОТЫ САЛИЦИЛОВОЙ В КОМБИНИРОВАННЫХ ЛЕКАРСТВЕННЫХ ПРЕПАРАТАХ

Introduction. In quality control of pharmaceutical formulations, there are methodological approaches based on the preliminary separation of sample components and approaches without preliminary separation. The use of solid-phase extraction (SPE) allows to separate analytes during sample preparation, with the separation selectivity determining the analysis selectivity as a whole. Currently, sorbents that provide selectivity in interaction with the analyte are of interest. Various derivatives of cellulose are used to obtain selective sorbents: ethylcellulose, hydroxypropylmethylcellulose (hypromellose), carboxymethylcellulose, etc. On the pharmaceutical market of the Russian Federation, there are pharmaceutical formulations with the active pharmaceutical substance salicylic acid. The use of selective SPE for analysis is a promising direction of research in drug quality control. Aim. Evaluation of the metrological characteristics of benzocaine quantitation by UV-spectrophotometry in certain pharmaceutical formulations using hypromellose sorbent for solid-phase extraction. Material and methods. The sorbent using a method developed by us was obtained. Salicylic acid (FS.2.1.0634, Russia State Pharmacopoeia XV ed.) to form active binding sites in sorbent structure was applied. The sorbent structure is polycyanoacrylate matrix with hypromellose fragments, the surface area is 255.50 m2/g, pore volume is 0.1433 cm3, pore diameter is 5.32 nm. The sorption capacity of hypromellose sorbent for benzocaine was 12.2 ± 0.8 μg/g. The SPE technique is proposed. SPE includes the stages: conditioning, sample addition and step-by-step elution with purified water and sodium hydroxide solution 0.1 mol/L. The spectrophotometer SF-56 was used to measure the absorbance at an analytical wavelength of 286 nm. To evaluate the selectivity (specificity) of sample preparation during chromatographic (SPE) separation of pharmaceutical formulations, absorption spectra of eluates in the wavelength range of 200-400 nm were scanned. The salicylic acid identification in the eluate was carried out based on absorption peaks at 228 and 298 nm.The salicylic acid calibration curve in the range of 1-20 μg/ml. To determination the metrological characteristics of salicylic acid quantitation method, 16 parallel determinations of samples of each pharmaceutical formulation were made. Statistical processing of the experimental results and determination of the metrological characteristics evaluation of analytical technique were carried out in accordance with general pharmacopoeial monograph (OFS.1.1.0013 "Statistical processing of results of physical, physicochemical, and chemical tests"). Results. When realizing the salicylic acid quantitation technique for measuring absorbance, it is advisable to use only the first portions of eluate (a solution of sodium hydroxide) in a volume of 5-10 ml were obtained. The relative error in determining the average content of salicylic acid in the analyzed preparations ranged from 1.18 to 1.44% for methods with solid-phase extraction (SPE) and 1.86-2.62% in its absence. It has been shown that there is a statistically significant difference in reproducibility for all preparations, as indicated by F-criteria values ranging from 7.81 to 13.5, exceeding their tabulated values. For the method without solid-phase extraction (SPE), there was an overestimation of the salicylic acid content in all cases, increasing the systematic error on average by 87.5%. Conclusion. The possibility of using a selective hypromellose sorbent for sample preparation for salicylic acid quantitation determination in pharmaceutical formulations has been showed. The use of sorbent for analysis provides to systematic error decrement of salicylic acid quantitation by an average of 87.5%.

APPLICATION OF SELECTIVE HYPROMELLOSE SORBENT FOR THE BENZOCAINE QUANTITATION IN PHARMACEUTICAL FORMULATIONS

Introduction. In quality control of pharmaceutical formulations, there are methodological approaches based on the preliminary separation of sample components and approaches without preliminary separation. The use of solid-phase extraction (SPE) allows to separate analytes during sample prepa-ration, with the separation selectivity determining the analysis selectivity as a whole. Currently, sorbents that provide selectivity in interaction with the analyte are of interest. Aim. Evaluation of the metrological characteristics of benzocaine quantitation by UV-spectrophotometry in certain pharmaceutical formulations using hypromellose sorbent for solid-phase extraction. Material and methods. The sorbent using a method developed by us was obtained. Benzocaine (FS.2.1.0634, Russia State Pharmacopoeia XV ed.) to form active binding sites in sorbent structure was applied. The sorbent structure is polycyanoacrylate matrix with hypromellose fragments, the sur-face area is 255.50 m2/g, pore volume is 0.1433 cm3, pore diameter is 5.32 nm. The sorption capacity of hypromellose sorbent for benzocaine was 12.2±0.8 μg/g. The SPE technique is proposed. SPE includes the stages: conditioning, sample addition and step-by-step elution with purified water and hydrochloric acid solution 0.1 mol/L. The spectrophotometer SF-56 was used to measure the absorbance at an analytical wavelength of 286 nm. To evaluate the selectivity (specificity) of sample preparation during chromatographic (SPE) separation of pharmaceutical formulations, absorption spectra of eluates in the wavelength range of 200400 nm were scanned. The benzocaine identification in the eluate was carried out based on absorption peaks at 220 and 286 nm. The benzocaine calibration curve in the range of 120 μg/ml. To determination the metrological characteristics of benzocaine quan-titation method, 11 parallel determinations of samples of each pharmaceutical formulation were made. Results. When realizing the benzocaine quantitation technique for measuring absorbance, it is advisable to use only the first portions of eluate (a solu-tion of hydrochloric acid) in a volume of 510 ml were obtained. The relative error of benzocaine average concentration in the pharmaceutical formula-tions ranged from 1.28 to 1.34% for technique that included the SPE stage, and from 1.86 to 2.02% for technique that did not include it. A comparison of the modifications of the spectrophotometric quantitation of benzocaine in pharmaceutical formulation using hypromellose sorbent and technique without the SPE did not reveal a statistically significant difference in reproducibility. For the modification technique without SPE, an increase of benzo-caine concentration and in determination relative error by an average of 48.0% were observed. Conclusion. The possibility of using a selective hypromellose sorbent for sample preparation for benzocaine quantitation determination in pharmaceu-tical formulations has been showed. The use of a sorbent for SPE at the sample preparation stage reduces the systematic error of benzocaine spectro-photometry quantitation for the analyzed pharmaceutical formulations by an average of 79.2%.

Synthesis, molecular docking and pharmacological evaluations of novel naphthalene-pyrazoline hybrids as new orally active anti-inflammatory agents

The pyrazoline nucleus is widely present in diverse compounds as it serves as a valuable template for the discovery of new class of bioactive compounds. Various substituted 2-pyrazolines exhibit a spectrum of pharmacological activities, including antimicrobial, anti-inflammatory, analgesic, and antitumor properties. In view of this, we have synthesized a novel class of naphthalene-pyrazoline hybrids (PY1-PY20). All the synthesized hybrids were thoroughly characterized by means of standard spectroscopic techniques and tested for their in-vivo anti-inflammatory activity. Most of the compounds exhibited significant anti-inflammatory activity and in particular the compounds bearing electron withdrawing groups had potential in vivo activity. Further, in-silico molecular docking studies were carried out using the ‘Molegro Virtual Docker’ as a docking tool against the target proteins such as COX-1 (PDB ID: 1EQH) and COX-2 (PDB ID: 1PXX). The results revealed that the majority of pyrazolines form hydrogen interactions with key residues (Ser 530, Tyr 385, Tyr 355, Arg 120, Ser 353, Arg 83) in the active binding sites of COX-1 (1EQH) and COX-2 (1PXX). Notably, interactions with residues Tyr 355, Arg 120, Ser 530 for COX-1, and Tyr 355, Arg 120, Gln 192, Leu 352, Ser 353, His 90, Tyr 38, Ser 530, and Tyr 385 for COX-2 are crucial for the reported anti-inflammatory activity of diverse compounds. Ligand-protein inverse molecular docking studies highlight specific hydrogen bond interactions, with PY-8, PY-11, PY-15, and PY-17 showing significant interactions. The study was able to identify potential anti-inflammatory naphthalene-pyrazoline derivatives.

Secondary metabolites of Trichoderma spp. as EGFR tyrosine kinase inhibitors: Evaluation of anticancer efficacy through computational approach.

The present study explores the epidermal growth factor receptor (EGFR) tyrosine kinase inhibition efficacy of secondary metabolites in Trichoderma spp. through molecular docking, molecular dynamics (MD) simulation and MM-PBSA approach. The result of molecular docking confirmed that out of 200 metabolites screened, three metabolites such as Harzianelactone A, Pretrichodermamide G and Aspochalasin M, potentially bound with the active binding site of EGFR tyrosine kinase domain(PDB ID: 1M17) with a threshold docking score of ≤- 9.0 kcal/mol when compared with the standard EGFR inhibitor (Erlotinib). The MD simulation was run to investigate the potential for stable complex formation in EGFR tyrosine kinase domain-unbound/lead metabolite (Aspochalasin M)-bound/standard inhibitor (Erlotinib)-bound complex. The MD simulation analysis at 100 ns revealed that Aspochalasin M formed the stable complex with EGFR. Besides, the in silico predication of pharmacokinetic properties further confirmed that Aspochalasin M qualified the drug-likeness rules with no harmful side effects (viz., hERG toxicity, hepatotoxicity and skin sensitization), non-mutagenicity and favourable logBB value. Moreover, the BOILED-Egg model predicted that Aspochalasin M showed a higher gastrointestinal absorption with improved bioavailability when administered orally and removed from the central nervous system (CNS). The results of the computational studies concluded that Aspochalasin M possessed significant efficacy in binding EGFR's active sites compared to the known standard inhibitor (Erlotinib). Therefore, Aspochalasin M can be used as a possible anticancer drug candidate and further in vitro and in vivo experimental validation of Aspochalasin M of Trichoderma spp. are required to determine its anticancer potential.

Characterizing the Molecular Mechanism of the Lethal C423D Mutation in FgMyoI: A Molecular Perspective.

The lethal mutation C423D in Fusarium graminearum myosin I (FgMyoI) occurs close to the binding pocket of the allosteric inhibitor phenamacril and causes severe inhibition on mycelial growth of F. graminearum strain PH-1. Here, based on extensive Gaussian accelerated molecular dynamics simulations and wet experiments, we elucidate the underlying molecular mechanism of the abnormal functioning of the FgMyoIC423D mutant at the atomistic level. Our results suggest that the damaging mutation C423D exhibits a synergistic allosteric inhibition mechanism similar to but more robust than that of phenamacril, including effects on the active site and actin binding. Unlike phenamacril-induced closure of Switch2, the mutation results in unfolding of the N-terminal relay helix with a partially opened Switch2 and blocks the structural rearrangement of the relay/SH1 helices, impairing the proper initiation of the recovery stroke. Due to the significant influence of C423D mutation on the function of FgMyoI, designing covalent inhibitors targeting this site holds tremendous potential.

Adrenodoxin allosterically alters human cytochrome P450 11B enzymes to accelerate substrate binding and decelerate release.

Two human mitochondrial membrane CYP11B enzymes play a pivotal role in steroidogenesis. CYP11B1 generates the major glucocorticoid cortisol, while CYP11B2 catalysis yields the primary mineralocorticoid aldosterone. Catalysis by both requires electron delivery by a soluble iron-sulfur adrenodoxin redox partner. However recent studies have shown that adrenodoxin/CYP11B interaction alone allosterically increases substrate and inhibitor affinity as exhibited by decreased dissociation constant (K d) values. The current study moves beyond such equilibrium studies, by defining adrenodoxin effects on the rates of P450 ligand binding and release separately. Stopped-flow data clearly demonstrate that adrenodoxin interaction with the P450 proximal surfaces increases ligand binding in both P450 CYP11B active sites by increasing the on rate constant and decreasing the off rate constant. As substrate entry and exit from the sequestered P450 active site requires conformational changes on the distal side of the P450 enzyme, a likely explanation is that adrenodoxin binding allosterically modulates CYP11B conformational changes. The 93% identical CYP11B enzymes can bind and hydroxylate each other's native substrates differing only by a hydroxyl. However, CYP11B1 exhibits monophasic substrate binding and CYP11B2 biphasic substrate binding, even when the substrates are swapped. This indicates that small differences in amino acid sequence between human CYP11B1 and CYP11B2 enzymes are more functionally important in ligand binding and could suggest avenues for more selective inhibition of these drug targets. Both protein/protein interactions and protein/substrate interactions are most likely to act by modulating CYP11B conformational dynamics.

PPX/PXP-type ligands (X = O and S) and their transition metal complexes: synthesis, properties and applications.

Short-bite diphosphines of the form R2P-X-PR2 (PXP; X = O, S; R = aryl, alkyl), incorporating an oxygen or sulphur atom as bridging unit X, are widely underexplored compared to their N- and C-containing PNP- and PCP-type counterparts. However, these PXP ligands undergo an interesting phosphorotropic equilibrium with the PPX (R2P(X)-PR2) tautomer, which opens up a very versatile coordination chemistry. This article covers the impact of the ligand backbone in short-bite ligands on their coordination chemistry, reactivity and applications. Especially in PXP-type complexes, metallophillic interactions can be induced in the case of coinage metals, which lead to fascinating photo-optical properties. Furthermore, PPX/PXP-type complexes are believed to exhibit a promising behavior in catalysis, due to the potential hemilability of the ligand and the therewith involved availability of free active sites for substrate binding.

Targeted suppression of oral squamous cell carcinoma by pyrimidine-tethered quinoxaline derivatives.

Oral cancer (OC) stands as a prominent cause of global mortality. Despite numerous efforts in recent decades, the efficacy of novel therapies to extend the lifespan of OC patients remains disappointingly low. Consequently, the demand for innovative therapeutic agents has become all the more pressing. In this context, we present our work on the design and synthesis of twenty-five novel quinoxaline-tethered imidazopyri(mi)dine derivatives. This was followed by comprehensive investigations into the impact of these molecules on the OC cell line. The in vitro cytotoxicity studies performed in CAL-27 and normal oral epithelial (NOE) cell lines revealed that some of the synthesized molecules like 12d have potent antiproliferative activity specifically towards OC cells with an IC50 of 0.79 μM and show negligible cytotoxicity over NOE cells. Further, 12d arrested cell growth in the S phase of the cell cycle and induced cell death by early apoptosis. The in silico studies validated that 12d binds to the activator binding site on pyruvate kinase M2 (PKM2) overexpressed in OC while the lactate dehydrogenase (LDH)-coupled enzyme assay established 12d as a potent PKM2 activator with an AC50 of 0.6 nM. Hence, this study provides fruitful evidence for the designed compounds as anticancer agents against OC.

Novel sulfonamides unveiled as potent anti-lung cancer agents via tumor pyruvate kinase M2 activation.

This rational pursuit led to the identification of a novel sulfonamide derivative as a potent anti-lung cancer (LC) compound. Considering these results, we synthesized 38 novel sulfonamide derivatives with diverse skeletal structures. In vitro cytotoxicity assays revealed a potent and selective antiproliferative effect against A549 cells after evaluating a panel of cancer cell lines. Compound 9b has emerged as a potent activator of tumor pyruvate kinase M2 (PKM2), a protein known to play a critical role in LC. Apoptosis assays and cell cycle analysis demonstrated early apoptosis and G2 phase arrest. In silico studies demonstrated interactions between compound 9b and the activator binding site of PKM2. Surface plasmon resonance (SPR) experiments strongly indicated that 9b has a high affinity (K d of 1.378 nM) for PKM2. Furthermore, the increase in reactive oxygen species and decrease in lactate concentration suggested that compound 9b has significant anticancer effects. Notably, the increase in particle size following treatment with 9b suggested the tetramerization of PKM2. This work provides insights that might advance efforts to develop effective non-platinum anticancer agents.

Hepatocellular Carcinoma Detection by Cell Sensor Based on Anti-GPC3 Single-Chain Variable Fragment

Precise identification of hepatocellular carcinoma (HCC) in early stages remains both a challenge and an opportunity for prevention and diagnosis of early liver cancer. Fast-detecting techniques like electrochemical sensing might provide more insight than current clinical methods owing to its high sensitivity, rapid detection, and convenient operation. Here, a single-chain variable fragment (scFv) was designed by retaining the antigen binding active sites of a recombinant humanized monoclonal antibody and modifying with histidine tags, and consequently used for the construction of cell sensor for HCC detection. The customized scFv showed specific binding signature on the membrane of HCC cell and positive correlation with cell concentration using fluorescence confocal microscopy and flow cytometry, respectively. A higher binding affinity of scFv over the original monoclonal antibody was also displayed. After the immobilization onto the screen-printed electrode through the high affinity between histidine and nickel-based nanomaterial, the constructed cell sensor exhibited a wide detection range of 10 2 to 10 7 cells ml −1 and a low detection limit of 2 cells ml −1 with a short incubation time of 5 min. The high selectivity in the liver tissue extract fluid derived from the design strategy was also displayed in the mouse samples. The customized scFv and related cell sensor demonstrated great potential for fast HCC analysis in early-stage cancer diagnosis.