Strong Hydrophobic Interactions Research Articles

Target-triggered double fluorescent biosensors for rapid and sensitive detection of long-chain perfluorinated compounds using DNA probe and lysozyme fiber

Perfluorinated compounds (PFCs) are useful man-made chemicals and serve as new emerging organic pollutants due to their environmental and health concerns. Chromatography-mass detection methods often need complex procedure and are also too expensive, so there is a critical demand to develop rapid, inexpensive, easy-to-operate and sensitive methods for PFCs detection. In this work, double fluorescent biosensors (‘DT sensor’ and ‘FT sensor’) have been designed to quantitatively detect long-chain perfluorinated compounds (PFCs), due to their strong hydrophobic interaction with DNA probe or lysozyme fiber. The ratio and rapid fluorescence responses offered more obvious signal changes, and high sensitivity with a limit of detection (LOD) of 0.16 μM (98.2 ppb) for perfluorododecanoic acid (PFDoA). For three PFCs with longer perfluoroalkyl chain (CF2), increased detection sensitivity was achieved due to a stronger hydrophobicity. The fluorescent biosensors showed a good selectivity for long-chain PFCs and served as cross-reactive sensors to differentiate three different long-chain PFCs. The biosensors also had robust signal response in tap water or serum samples, and the LOD can be further lowered to pM (ppt) level after sample preconcentration.

Dynamic Interactions Between Brilliant Green and MscL Investigated by Solid-State NMR Spectroscopy and Molecular Dynamics Simulations.

The mechanosensitive ion channel of large conductance (MscL) is a promising template for the development of new antibiotics due to its high conservation and uniqueness to microbes. Brilliant green (BG), a triarylmethane dye, has been identified as a new antibiotic targeted MscL. However, the detailed binding sites to MscL and the dynamic pathway of BG through the MscL channel remain unknown. Here, the dynamic interactions between BG and MscL were investigated using solid-state NMR spectroscopy and molecule dynamics (MD) simulations. Residue site-specific binding sites of BG to the MscL channel were identified by solid-state NMR. In addition, MD simulations revealed that BG conducts through the MscL channel via residues along the inner surface of the pore sequentially, in which the strong hydrophobic interactions between BG and hydrophobic residues F23 and I27 in the hydrophobic gate region of the MscL channel are major restrictions. Particularly, it was demonstrated that BG activates the MscL channel by reducing the hydrophobicity of the F23 in the gate region by water molecules that are bound to BG. Taken together, these simulations and experimental data provide novel insights into the dynamic interactions between BG and MscL, based on which new hydrophobic antibiotics and adjuvants targeting MscL can be developed.

Improved Light and In Vitro Digestive Stability of Lutein-Loaded Nanoparticles Based on Soy Protein Hydrolysates via Pepsin.

In order to improve the water solubility and stability of lutein, soy protein isolates (SPI) and their hydrolysates via pepsin (PSPI) and alcalase (ASPI) were used as nanocarriers for lutein to fabricate the lutein-loaded nanoparticles (LNPS) of SPI, PSPI, and ASPI. The encapsulation properties, light, and in vitro digestive stability of lutein in nanoparticles, and protein-lutein interactions were investigated. Compared with SPI-LNPS and ASPI-LNPS, PSPI-LNPS was characterized by uniform morphology (approximately 115 nm) with a lower polydispersity index (approximately 0.11) and higher lutein loading capacity (17.96 μg/mg protein). In addition, PSPI-LNPS presented the higher lutein retention rate after light exposure (85.05%) and simulated digestion (77.73%) than the unencapsulated lutein and SPI-LNPS. Fluorescence spectroscopy revealed that PSPI had stronger hydrophobic interaction with lutein than SPI, which positively correlated with their beneficial effects on the light and digestive stability of lutein. This study demonstrated that PSPI possessed significant potential for lutein delivery.

Insights into the Inhibitory Mechanism of Viniferifuran on Xanthine Oxidase by Multiple Spectroscopic Techniques and Molecular Docking.

Viniferifuran was investigated for its potential to inhibit the activity of xanthine oxidase (XO), a key enzyme catalyzing xanthine to uric acid. An enzyme kinetics analysis showed that viniferifuran possessed a strong inhibition on XO in a typical anti-competitive manner with an IC50 value of 12.32 μM (IC50 for the first-line clinical drug allopurinol: 29.72 μM). FT-IR and CD data analyses showed that viniferifuran could induce a conformational change of XO with a decrease in the α-helix and increases in the β-sheet, β-turn, and random coil structures. A molecular docking analysis revealed that viniferifuran bound to the amino acid residues located within the activity cavity of XO by a strong hydrophobic interaction (for Ser1214, Val1011, Phe914, Phe1009, Leu1014, and Phe649) and hydrogen bonding (for Asn768, Ser876, and Tyr735). These findings suggested that viniferifuran might be a promising XO inhibitor with a favorable mechanism of action.

Metal-Ion-Induced Evolution of Phenylalanine Self-Assembly: Structural Polymorphism of Novel Metastable Intermediates.

The self-assembly of aromatic amino acids has been widely studied due to their ability to form well-defined amyloid-like fibrillar structures. Herein, for the first time, we report the existence of different metastable intermediate states of diverse morphologies, for example, droplets, spheres, vesicles, flowers, and toroids, that are sequentially formed in aqueous medium during the self-assembly process of phenylalanine in the presence of different divalent (Zn2+, Cd2+, and Hg2+) and trivalent (Al3+, Ga3+, and In3+) metal ions having low pKa values. Due to metal ion-amino acid coordination and strong hydrophobic interaction induced by these metal ions, spherical aggregates are obtained at the initial stage of the structural evolution and further transformed into other intermediate states. Our work may facilitate understanding of the role of metal ions in the amino acid self-assembly process and broaden future applications of the obtained nanostructures in drug delivery, tissue engineering, bioimaging, biocatalysis, and other fields.

β-Carboline dimers inhibit the tumor proliferation by the cell cycle arrest of sarcoma through intercalating to Cyclin-A2.

β-Carbolines are potentially strong alkaloids with a wide range of bioactivities, and their dimers exhibit stronger antitumor activity other than the monomers. However, the detailed mechanisms of the β-carboline dimers in inhibiting sarcoma (SARC) remain unclear. The results showed that β-carboline-3-carboxylic acid dimers Comp1 and Comp2, which were synthesized in our lab and modified at the N9 position and linked at the C3 position, exhibited effective inhibition activity on MG-63 proliferation (IC50 = 4.6μM). Meanwhile, the large scale transcriptome profiles of SARC from The Cancer Genome Atlas (TCGA) were analyzed, and found that abnormal expression of genes relevant to apoptosis, cell cycle, and signaling pathways of Hedgehog, HIF, Ras involved in the SARC pathogenesis. Interestingly, both dimers could promote the apoptosis and arrest the cell cycle in S phase to inhibit proliferation of MG-63. Moreover, Comp1 and Comp2 inhibited the expression CDK2, CCNA2, DBF4, and PLK1 associated with various immune cells and cell cycle in MG-63. Remarkably, drug-target interaction network analysis showed that numerous proteins involved in cell cycle were the potential targets of Comp1 and Comp2, especially CCNA2. Further molecular docking, isothermal titration calorimetry (ITC) and Cellular Thermal Shift Assay (CETSA) confirmed that both dimers could directly interact with CCNA2, which is significantly correlated with CD4+ T cells, by strong hydrophobic interactions (Kd=5.821 ×106 N). Meanwhile, the levels of CCNA2 and CDK2 were inhibited to decrease in MG-63 by both dimer treatments at transcription and protein levels, implying that Comp1 and Comp2 blocked the interaction between CCNA2 and CDK2 through competitive binding with CCNA2 to arrest the cell cycle of MG-63 cells in the S phase. Additionally, the transcriptome profiles of β-carboline-treated mice from Gene Expression Omnibus (GEO) were obtained, and found that similar antitumor mechanism was shared among β-carboline derivatives. Overall, our results elucidated the antitumor mechanisms of Comp1 and Comp2 through dual-suppressing the function of CCNA2 to profoundly arrest cell cycle of MG-63, then effectively inhibited cell proliferation of MG-63. These results provide new insights into the antitumor mechanism of β-carboline dimers and new routes of various novel cancer-related drug targets for future possible cancer therapy.

Luminescent Gold Nanoparticles with Controllable Hydrophobic Interactions.

The construction of luminescent gold nanoparticles (AuNPs) with highly redshifted emission in the second near-infrared window (NIR-II) and good biocompatibility is still challenging. Herein, using an amphiphilic block copolymer (ABC) template with controllable hydrophobic interactions in the diverse forms of unimers and micelles, we report a facile strategy for redshifting the emission and enhancing the biological interactions of luminescent AuNPs. While the uniform clusters of NIR-II AuNPs are formed in situ inside the hydrophobic cores of ABC micelles with strong interparticle hydrophobic interactions and enhanced emission at 1080 nm with a high quantum yield (QY) of 1.6%, the rigid NIR-II AuNPs are generated with strong intraparticle hydrophobic interactions as ABC unimers on the surface, leading to a redshifted emission of 1280 nm with a QY of 0.25% and enhancing the affinities toward injured intestinal mucosa in colitis imaging. These findings open new possibilities for the design of highly redshifted luminescent AuNPs with enhanced biological interactions.

(Digital Presentation) A Coarse-Grained Molecular Dynamics Study on the Aggregation and Adsorption Behavior of Ionomer from Solution Onto Pt/C Substrate

The catalyst layer (CL) was an important component in polymer electrolyte fuel cells (PEFCs). In the CL, Nafion ionomers coated the Pt particles on the carbon surface for the proton transfer assistance, and therefore the structure optimization of ionomer films is very important for the improvement of catalytic utilization and proton transport performance. The microscopic structure of ionomer covered carbon and Pt particles in the solution was related to the final structural characteristics of the ionomer film determining the performance of CL. In order to elucidate the structural evolution and drying process of the catalyst ink, in the present study, we carried out coarse-grained molecular dynamics (CGMD) simulations to investigate the adsorption behavior of ionomers and morphology evolution of Nafion ionomer from the aqueous solutions to the Pt/C substrate surface under various solution conditions including ionomer concentration and I/C ratio, and substrate conditions including the size and distribution of Pt particles, and Pt/C ratio. We found that the ionomers formed a cylindrical aggregate in the solution and are easily attracted by Pt particles due to their hydrophilic ionic shells. Although ionomer aggregates are easily to adsorb onto Pt particles, it tended to unfold and spread onto the carbon substrate finally, which is resulted from the hydrophobic attraction of the exposed ionomer backbone to the carbon surface and the hydrophilic attraction of ionomer sulfonate groups to the aqueous solution. The ionomer adsorption displayed a strong correlation with the ionomer concentration in the dilute solution. However, the I/C ratio had little effect on ionomer adsorption. This adsorption phenomenon of ionomers in the dilution solution was found to fit the Langmuir adsorption model very well but failed when the concentration of ionomer became higher. A wettability switching behavior of ionomers is observed because the morphology of ionomers changed from the solution to the surface. Two types of the adsorption behavior of ionomer aggregates in the aqueous solution were observed. The first adsorption behavior was the direct adsorption of ionomer aggregate to the carbon substrate without any interaction with Pt particles. For the ionomer aggregate, exposed apolar backbones of ionomers that were close to the carbon could help the ionomer aggregate adsorb on the hydrophobic carbon surface. During the adsorption process, cylindrical-like shape ionomer aggregates gradually spread onto the carbon surface. Finally, the ionomers were entirely distributed on the carbon surface without aggregating together. The second adsorption behavior was indirect adsorption of ionomer aggregate, where the ionomer aggregates firstly adsorbed onto the Pt particles from solution, and finally to the carbon substrate. In the solution, the apolar backbones of ionomer aggregates located at the core of the aggregate were difficult to contact with the substance outside such as carbon surface, while the anionic groups located at the aggregate-solvent interface were easy to contact with the substance outside. When such Nafion aggregates were close to the Pt particles, the anionic shells of the cylindrical aggregates were easily to adsorb onto hydrophilic Pt particles. When the Nafion aggregate adsorbed onto Pt particles, Nafion still presents a cylinder-like aggregate state until it started to approach the carbon surface. When the Nafion aggregate begin to move onto the carbon substrate, it gradually unfolded due to the strong hydrophobic interaction between the ionomer backbone and the carbon surface. During this process, Nafion ionomers gradually relieved their internal hydrophobic associations between their backbones because the unfolded ionomers created more surface contacts and better anchored themselves to the surface. Therefore, when the exposed carbon surface was large enough, increasing the number and sizes of Pt particles could increase the surface area of Pt particles, which favors the ionomer adsorption. When the space between Pt particles was too small to create enough carbon surface contacts anchoring the ionomer backbone, the coverage of the ionomer dropped rapidly. The present study provided insight into ionomer adsorption from solutions to Pt/C surface, which could contribute to the catalyst ink design to improve the structure of catalyst layers.AcknowledgmentsThis work was supported by the New Energy and Industrial Technology Development Organization (NEDO) of Japan, Grant number JPNP20003. It was performed on the Supercomputer system “AFI-NITY” at the Advanced Fluid Information Research Center, Institute of Fluid Science, Tohoku University. We also thank W. Shirakawa for her assistance with the simulations.

Synthesis of linear and cyclic organic sulfonic acid-modified Cu-Al layered double hydroxides and their adsorption properties

Over the past few years, many substances have been investigated for their potential application as adsorbent materials. The combination of a layered double hydroxide (LDH) and CH/π interactions enables selective adsorption through the differences in the electron density of the aromatic ring. Thus, in this study, two organic anions, namely the linear 1-octane sulfonate (OS-) and cyclic sulfonated-β-cyclodextrin (SCD13-), were intercalated into the interlayers of a Cu-Al LDH by coprecipitation (Cu/Al ratio, 3.0; chemical ratio, 2.0). The SCD13- anion was selected because β-cyclodextrin comprises hydrophobic cavities that allow the formation of inclusion compounds. X-ray diffraction (XRD), total organic carbon (TOC) analysis, and inductively coupled plasma-atomic emission spectrometry (ICP-AES) revealed that the as-synthesized OS-Cu-Al LDH and SCD-Cu-Al LDH were both oriented perpendicular to the LDH layer. Subsequent absorption experiments with m-aminophenol (AP) and m-nitrophenol (NP), which have similar structures but different aromatic ring electron densities, confirmed a Freundlich-type adsorption mechanism and revealed that adsorption occurred on a non-uniform surface for both LDHs. The results also revealed excellent NP and AP adsorptions for the linear OS•Cu-Al LDH and cyclic SCD•Cu-Al LDH, respectively. This was attributed to the strong hydrophobic interactions in the former and strong CH/π interactions in the latter. This study shows the potential of using linear and cyclic organic sulfonic acid-modified LDHs for the selective recovery of the desired aromatic organic compounds.

Self-Assembly Anchored Cationic Copolymer Interfaces for Applying the Control of Counterion-Induced Bacteria Killing/Release Procedure.

In recent years, daily hygiene and disease control issues have received increasing attention, especially the raging epidemics caused by the spread of deadly viruses. The construction of the interface of new polymer materials is focused on, which can provide a cyclic operation process for the killing and releasing of bacteria, and perform repeated regeneration, which is of great significance for the development of advanced medical biomaterials. In order to explore the basic physical phenomena of bacterial attachment and detachment on the polymer material interface by different amine groups, this study plans to synthesize four different butyl methacrylate (BMA)-based cationic copolymers with primary, ternary, and quaternary amine groups, and compare their effects on bactericidal efficiency. Since BMA can generate strong hydrophobic interactions with the benzene ring structure, this study used a polystyrene substrate to realize a self-assembled cationic copolymer interface for controlling the counterion-induced bacterial killing/release process. Furthermore, negatively charged ions are introduced to induce changes in the hydration capability of water molecules and control the subsequent bacterial detachment function. In this study, possible directions to answer and clarify the above concepts are proposed, and there is a basic reference principle that can lead to research work in macromolecular biosciencefields.

Charge-Convertible and Reduction-Sensitive Cholesterol-Containing Amphiphilic Copolymers for Improved Doxorubicin Delivery.

For achieving successful chemotherapy against cancer, designing biocompatible drug delivery systems (DDSs) with long circulation times, high cellular endocytosis efficiency, and targeted drug release is of upmost importance. Herein, a well-defined PEG-b-P(MASSChol-co-MANBoc) block copolymer bearing redox-sensitive cholesteryl-side group was prepared via reversible addition-fragmentation chain transfer (RAFT) polymerization (with non-redox PEG-b-P(MACCChol-co-MAN-DCA) as the reference), and 1,2-dicarboxylic-cyclohexene acid (DCA) was then grafted onto the hydrophobic block to endow it with charge-convertible characteristics under a tumor microenvironment. The amphiphilic copolymer could be assembled into polymeric spherical micelles (SSMCs) with polyethylene glycol (PEG) as the corona/shell, and anti-cancer drug doxorubicin (DOX) was successfully encapsulated into the micellar core via strong hydrophobic and electrostatic interactions. This nanocarrier showed high stability in the physiological environment and demonstrated “smart” surface charge conversion from negative to positive in the slightly acidic environment of tumor tissues (pH 6.5~6.8), as determined by dynamic light scattering (DLS). Moreover, the cleavage of a disulfide bond linking the cholesterol grafts under an intracellular redox environment (10 mM GSH) resulted in micellar dissociation and accelerated drug release, with the non-redox-responsive micelles (CCMCs) as the control. Additionally, a cellular endocytosis and tumor proliferation inhibition study against MCF-7 tumor cells demonstrated the enhanced endocytosis and tumor cell inhibitory efficiency of dual-responsive SSMCs/DOX nanomedicines, revealing potentials as multifunctional nanoplatforms for effective oncology treatment.

Experimental Study on Strength and Microstructure of Glacial Till Stabilized by Ionic Soil Stabilizer

Glacial till, widely distributed in southwest China, is a special soil directly deposited by detritus formed from melting Quaternary glaciers. In this paper, the F1 ionic soil stabilizer was adopted for stabilizing the glacial till to improve its mechanical strength. A series of micro and macro tests were carried out to study the mechanical properties and microstructure of stabilized soil with the F1 ionic soil stabilizer. The results show that the F1 ionic soil stabilizer can destroy the diffuse double layer structure on the surface of glacial till particles and reduce the thickness of the adsorbed water layer through strong cation exchange and hydrophobic interactions of active sulfonated oil, which reduce the spacing of glacial till particles, enhance the aggregation of glacial till particles, and effectively suppress the swelling deformation of the glacial till. It can be concluded that the water sensitivity and compaction characteristics of glacial till can be significantly improved by the stabilization of the F1 ionic soil stabilizer. Moreover, the mechanical strength of the glacial till can be significantly improved by the stabilization of the F1 ionic soil stabilizer. This article is helpful as a guideline for practical design and future research on applying the F1 ionic soil stabilizer to improve the bearing capacity of foundations in glacial till areas.

Long-term tracking of endoplasmic reticulum autophagy by a fluorescent probe with a long alkyl chain

Autophagy is a process through which cytoplasmic materials are degraded in lysosome. It is critical in the maintenance of cellular homeostasis. Thus, visualization and tracking of the autophagy is very important for understanding complex intracellular events. To monitor autophagy, various specific fluorescent probes to organelles are often used, such as mitochondria probes for mitophagy, lipid droplets probes for lipophagy. However, an endoplasmic reticulum (ER) probe for ER-phagy is still lack and challenging. Herein, we presented an ER fluorescent probe (HTMP-16) with a long alkyl chain. By strong hydrophobic interaction between the long alkyl chain and the lipid bilayers in ER, HTMP-16 can exclusively locate in the ER for a long time. At the same time, HTMP-16 exhibits excellent self-delivery to living cell, which is a prerequisite of a tracking probe. Furthermore, utilizing HTMP-16, in-situ and real-time long-term tracking of ER-phagy has been realized in living cells. Besides, considering its high sensitivity to viscosity, HTMP-16 can visualize the viscous changes of ER induced by temperature changes or tunicamycin.

Polypropylene membranes with high adsorption capacity and anti-adhesion properties achieved by hydrophobic interactions and hydrogen bonded self-assembly for uranium extraction from seawater

Herein, we have developed a mild and convenient strategy to enable polypropylene (PP) fibrous membranes with abundant adsorption sites for the selective extraction of uranium from seawater. Lysozyme was chosen as the pre-coatings because of its strong hydrophobic interactions with PP membranes. Then, lysozyme and phytic acid were set as hydrogen bonding self-assembling components since both of them are proved with a great binding affinity towards uranium. The contents and thickness of the lysozyme-phytic acid coatings could be linearly tuned due to the excellent controllability of self-assembly. The obtained PP membrane could achieve adsorption capacity as high as 90.07 mg-U/g-membrane (2143.83 mg-U/g-coating). In addition, the lysozyme would endow the PP membranes with outstanding anti-fouling, anti-bacteria, and anti-adhesion properties. The modified membranes could maintain more than 85% of adsorption capacity even with the interferences of bacteria and model pollutants. We also applied the membranes for dynamic adsorption by taking advantage of membrane filtrations. The adsorption capacity during the dynamic process increases with the flow rates and the layer numbers because more uranyl ions could contact with the adsorption sites. The modified PP membranes could obtain 1.15 mg/g of uranium after 15 days when the concentration of uranium is set as the same as the uranium concentration (3.3 μg/L) in natural seawater.

Fabrication and characterization of l-ascorbyl palmitate and phospholipid-based hybrid liposomes and their impacts on the stability of loaded hydrophobic polyphenols

The aim of this study was to evaluate the influence of l-ascorbyl palmitate (LAP) as an additive to liposome formulations by self-assembling with soy lecithin to form hybrid liposomes, in order to enhance the physical stability and bioactivator-loaded retention ratio of the LAP incorporated liposomes (LAP-LP). The addition of LAP significantly increased its surface negative charge and strong hydrophobic interactions occurred between the hydrophobic tails of LAP and phospholipids resulting in more compactly ordered, rigid and hydrophobic phospholipid bilayers as indicated by surface tension, fluorescence probes and DSC. These changes enhanced the stability of hydrophobic polyphenol loaded LAP-LP during storage. Particularly, after four weeks storage at 37 °C for naringenin loaded liposomes, the retention ratio of pure liposome decreased dramatically to 12.5 %, while the LAP-LP remained above 74.5 %. This study opens up the potential for the LAP-LP to be developed as a food-grade multifunctional formulation for encapsulating and delivering bioactivators.

Effect of Alkyl Side Chain Length on the Lithium-Ion Conductivity for Polyether Electrolytes.

The design guidelines of polymer structure to effectively promote lithium-ion conduction within the polymer electrolytes (PEs) are crucial for its practical use. In this study, the electrolyte properties of a simple polyether having alkyl side chains with varied lengths (−(CH2)m−H, m = 1, 2, 4, 6, 8, and 12) were compared and established a valid design strategy based on the properties of the alkyl side chain. Various spectro-electrochemical measurements successfully connected the electrolyte properties and the alkyl side chain length. Steric hindrance of the alkyl side chain effectively suppressed the interaction between ether oxygen and lithium-ion (m ≥ 2), decreasing the glass transition temperature and the activation energy of lithium-ion transfer at the electrode-electrolyte interface. The strong hydrophobic interactions aligned and/or aggregated the extended alkyl group (m ≥ 8), creating a rapid lithium-ion transport pathway and enhancing lithium-ion conductivity. A clear trend was observed for the following three crucial factors determining bulk lithium-ion transport properties along with the extension of the alkyl side chain: 1) salt dissociability decreased due to the non-polarity of the alkyl side chain, 2) segmental mobility of polymer chains increased due to the internal plasticizing effect, and 3) lithium-ion transference number increased due to the inhibition of the bulky anion transport by its steric hindrance. The highest lithium-ion conductivity was confirmed for the PEs with an alkyl side chain of moderate length (m = 4) at 70°C, indicating the optimized balance between salt dissociability, polymer segmental mobility, and selective lithium-ion transfer. The length of an alkyl side chain can thus be a critical factor in improving the performance of PEs, including thermal stability and lithium-ion conductivity. Precise tuning of the alkyl side chain-related parameters such as steric hindrance, polarity, internal plasticizing effect, and self-alignment optimizes the polymer segmental mobility and salt dissociability, which is crucial for realizing high lithium-ion conductivity for PEs.

Key interactions of pyrimethamine derivatives specific to wild-type and mutant P. falciparum dihydrofolate reductase based on 3D-QSAR, MD simulations and quantum chemical calculations

Plasmodium falciparum dihydrofolate reductase-thymidylate synthase (PfDHFR-TS) is an important target enzyme in malarial chemotherapy. An understanding of how novel inhibitors interact with wild-type (wtPfDHFR), quadruple-mutant (qmPfDHFR), and human (hDHFR) enzymes is required for the development of these compounds as antimalarials. This study is focused on a series of des-Cl and m-Cl phenyl analogs of pyrimethamine with various flexible 6-substituents. The interactions of these compounds with DHFR enzymes were investigated by 3 D-QSAR, MD simulations, MM-PBSA, and DFT calculations. CoMFA and CoMSIA models were developed with good predictive abilities for wtPfDHFR and qmPfDHFR. For hDHFR, CoMSIA models combined with clogP descriptor were successfully derived. Binding free energy using MM-PBSA and comparison of per residue decomposition energy analyses with the DFT method at M06-2X/6-31G ++(d,p) level of theory indicated that Asp54 and Phe58 play important roles in the binding of the most potent compound in the series (compound 27) with both wtPfDHFR and qmPfDHFR, whereas Arg59 and Arg122 were additionally found to interact with this inhibitor in qmPfDHFR. For hDHFR, the residues Glu30 and Phe34 but not Arg70, equivalent to Asp54, Phe58, and Arg122 in PfDHFR, also play role in compound 27 binding through strong hydrophobic interactions (Phe34) and hydrogen bond network with Glu30, Ile7, and Val115. From the key interactions identified in the DHFR-inhibitor complexes, a general scheme is proposed for designing new inhibitors selective for PfDHFR that is important for the development of novel antifolate antimalarials. Communicated by Ramaswamy H. Sarma

A systematic exploration of boceprevir-based main protease inhibitors as SARS-CoV-2 antivirals

Boceprevir is an HCV NSP3 inhibitor that was explored as a repurposed drug for COVID-19. It inhibits the SARS-CoV-2 main protease (MPro) and contains an α-ketoamide warhead, a P1 β-cyclobutylalanyl moiety, a P2 dimethylcyclopropylproline, a P3 tert-butylglycine, and a P4 N-terminal tert-butylcarbamide. By introducing modifications at all four positions, we synthesized 20 boceprevir-based MPro inhibitors including PF-07321332 and characterized their MPro inhibition potency in test tubes (in vitro) and 293T cells (in cellulo). Crystal structures of MPro bound with 10 inhibitors and cytotoxicity and antiviral potency of 4 inhibitors were characterized as well. Replacing the P1 site with a β-(S-2-oxopyrrolidin-3-yl)-alanyl (Opal) residue and the warhead with an aldehyde leads to high in vitro potency. The original moieties at P2, P3 and the P4 N-terminal cap positions in boceprevir are better than other tested chemical moieties for high in vitro potency. In crystal structures, all inhibitors form a covalent adduct with the MPro active site cysteine. The P1 Opal residue, P2 dimethylcyclopropylproline and P4 N-terminal tert-butylcarbamide make strong hydrophobic interactions with MPro, explaining high in vitro potency of inhibitors that contain these moieties. A unique observation was made with an inhibitor that contains a P4 N-terminal isovaleramide. In its MPro complex structure, the P4 N-terminal isovaleramide is tucked deep in a small pocket of MPro that originally recognizes a P4 alanine side chain in a substrate. Although all inhibitors show high in vitro potency, they have drastically different in cellulo potency to inhibit ectopically expressed MPro in human 293T cells. In general, inhibitors with a P4 N-terminal carbamide or amide have low in cellulo potency. This trend is reversed when the P4 N-terminal cap is changed to a carbamate. The installation of a P3 O-tert-butyl-threonine improves in cellulo potency. Three molecules that contain a P4 N-terminal carbamate were advanced to cytotoxicity tests on 293T cells and antiviral potency tests on three SARS-CoV-2 variants. They all have relatively low cytotoxicity and high antiviral potency with EC50 values around 1 μM. A control compound with a nitrile warhead and a P4 N-terminal amide has undetectable antiviral potency. Based on all observations, we conclude that a P4 N-terminal carbamate in a boceprevir derivative is key for high antiviral potency against SARS-CoV-2.

Molecular Mechanism of Interaction of Curcumin with BSA, Surfactants and Live E. Coli Cell Membrane Revealed by Fluorescence Spectroscopy and Confocal Microscopy.

Clinical trials on the therapeutic effect of curcumin have proven to be highly effective against many diseases including cancer and Alzheimer's. However, the molecular mechanism of interaction of curcumin with protein and live cell membrane is poorly understood. Here, we report the mechanism of interaction of curcumin with bovine serum albumin (BSA) and live E. coli cell membrane in the presence of organized assemblies of sodium dodecyl sulfate (SDS) and cetrimonium bromide (CTAB) by fluorescence spectroscopy, laser-scanning confocal microscopy, and computation. Enhanced binding constant, blue-shifted emission spectra, and imaging of heterogeneous FRET on live bacteria cell membrane strongly indicate the complex formation of curcumin with strong hydrophobic interaction, which is further validated by computation. Finally, our results may shed light on the efficient strategy of applications of curcumin as a natural therapeutic lead in clinical trials against many life-threatening diseases.

Apo-Form Selective Inhibition of IDO for Tumor Immunotherapy.

The pharmacological inhibition of IDO1 is considered an effective therapeutic approach for cancer treatment. However, the inadequate response of existing holo-IDO1 inhibitors and unclear biomarkers available in clinical practice limit the possibility of developing efficacious IDO1 inhibitors. In the current study, we aimed to elucidate the activity and mechanism of a potent 1H-pyrrole-2-carboxylic acid derivative (B37) targeting apo-IDO1 and to determine its role in tumor therapy. By competing with heme for binding to apo-IDO1, B37 potently inhibited IDO1 activity, with an IC50 of 22 pM assessed using a HeLa cell-based assay. The x-ray cocrystal structure of the inhibitor-enzyme complex showed that the B37-human IDO1 complex has strong hydrophobic interactions, which enhances its binding affinity, determined using isothermal titration calorimetry. Stronger noncovalent interactions, including π stacking and hydrogen bonds formed between B37 and apo-human IDO1, underlay the enthalpy-driven force for B37 for binding to the enzyme. These binding properties endowed B37 with potent antitumor efficacy, which was confirmed in a mouse colon cancer CT26 syngeneic model in BALB/c mice and in an azoxymethane/dextran sulfate sodium-induced colon carcinogenesis model in C57BL/6 mice by activating the host immune system. Moreover, the combination of B37 and anti-PD1 Ab synergistically inhibited tumor growth. These results suggested that B37 may serve as a unique candidate for apo-IDO1 inhibition-mediated tumor immunotherapy.