Natural products are widely used for treating mitochondrial dysfunction-related diseases and cancers. Curcumin, a well-known natural product, can be potentially used to treat cancer. Human salt-induced kinase 3 (SIK3) is one of the target proteins for curcumin. However, the interactions between curcumin and human SIK3 have not yet been investigated in detail. In this study, we studied the binding models for the interactions between curcumin and human SIK3 using computational tools such as homology modeling, molecular docking, molecular dynamics simulations, and binding free energy calculations. The open activity loop conformation of SIK3 with the ketoenol form of curcumin was the optimal binding model. The I72, V80, A93, Y144, A145, and L195 residues played a key role for curcumin binding with human SIK3. The interactions between curcumin and human SIK3 were also investigated using the kinase assay. Moreover, curcumin exhibited an IC50 (half-maximal inhibitory concentration) value of 131 nM, and it showed significant antiproliferative activities of 9.62 ± 0.33 µM and 72.37 ± 0.37 µM against the MCF-7 and MDA-MB-23 cell lines, respectively. This study provides detailed information on the binding of curcumin with human SIK3 and may facilitate the design of novel salt-inducible kinases inhibitors.

Lysophosphatidylserine (LysoPS) is an endogenous pan-agonist of three G-protein coupled receptors (GPCRs): LPS1/GPR34, LPS2/P2Y10, and LPS3/GPR174, and we previously reported a series of LysoPS-based agonists of these receptors. Interestingly, we found that LPS1 agonist activity was very sensitive to structural change at the hydrophobic fatty acid moiety, whereas LPS2 agonist activity was not. Here, to probe the molecular basis of LPS2 agonist binding, we developed a new class of hydrophobic fatty acid surrogates having a biphenyl-ether scaffold. The LPS2 agonist activity of these compounds proved sensitive to molecular modification of the hydrophobic skeleton. Thus, we next constructed an LPS2 model by homology modeling and docking/molecular dynamics (MD) simulation, and validated it by means of SAR studies together with point mutations of selected receptor amino-acid residues. The putative ligand-binding site of LPS2 is Γ-shaped, with a hydrophilic site horizontally embedded in the receptor transmembrane helix bundles and a perpendicular hydrophobic groove adjoining transmembrane domains 4 and 5 that is open to the membrane bilayer. The binding poses of LPS2 agonists to this site are consistent with easy incorporation of various kinds of fatty acid surrogates. Structural development based on this model afforded a series of potent and selective LPS2 full agonists, which showed enhanced in vitro actin stress fiber formation effect.

Microbial bioremediation is a very potent and eco-friendly approach to alleviate pesticide pollution in agricultural ecosystems, and hydrolase is an effective element for contaminant degradation. In the present study, a novel Mn2+-dependent esterase, PchA, that efficiently hydrolyzes carbamate pesticides with aromatic structures was identified from Pseudomonas sp. PS21. The hydrolytic activity was confirmed to be related closely to the core catalytic domain, which consists of six residues. The crucial residues indirectly stabilized the position of carbaryl via chelating Mn2+ according to the binding model clarified by molecular simulations, and the additional hydrophobic interactions between carbaryl with several hydrophobic residues also stabilized the binding conformation. The residue Glu398, by serving as the general base, might activate a water molecule and facilitate PchA catalysis. This work offers valuable insights into the binding interaction and hydrolytic mechanism of carbaryl with the hydrolase PchA and will be crucial to designing strategies leading to the protein variants that are capable of degrading related contaminants.

The rate of drug delivery to cells and the subsequent rate of drug metabolism are dependent on the cell membrane permeability to the drug. In some cases, tissue may be composed of different types of cells that exhibit order of magnitude differences in their membrane permeabilities. This paper presents a brief review of the components of the tissue scale three-compartment pharmacokinetic model of drug delivery to single-cell-type populations. The existing model is extended to consider tissue composed of two different cell types. A case study is presented of infusion mediated delivery of doxorubicin to a tumor that is composed a drug reactive cell type and of a drug resistive cell type. The membrane permeabilities of the two cell types differ by an order of magnitude. A parametric investigation of the population composition is conducted and it is shown that the drug metabolism of the low permeability cells are negatively influenced by the fraction of the tissue composed of the permeable drug reactive cells. This is because when the population is composed mostly of drug permeable cells, the extracellular space is rapidly depleted of the drug. This has two compounding effects: (i) locally there is simply less drug available to the neighboring drug resistant cells, and (ii) the depletion of the drug from the extracellular space near the vessel-tissue interface leaves less drug to be transported to booth cell types farther away from the vessel.

Destruxin A (DA), a hexa-cyclodepsipeptidic mycotoxin produced by the entomopathogenic fungus Metarhizium anisopliae, exhibits insecticidal activities in a wide range of pests and is known as an innate immunity inhibitor. However, its mechanism of action requires further investigation. In this research, the interactions of DA with the six aminoacyl tRNA synthetases (ARSs) of Bombyx mori, BmAlaRS, BmCysRS, BmMetRS, BmValRS, BmIleRS, and BmGluProRS, were analyzed. The six ARSs were expressed and purified. The BLI (biolayer interferometry) results indicated that DA binds these ARSs with the affinity indices (KD) of 10-4 to 10-5 M. The molecular docking suggested a similar interaction mode of DA with ARSs, whereby DA settled into a pocket through hydrogen bonds with Asn, Arg, His, Lys, and Tyr of ARSs. Furthermore, DA treatments decreased the contents of soluble protein and free amino acids in Bm12 cells, which suggested that DA impedes protein synthesis. Lastly, the ARSs in Bm12 cells were all downregulated by DA stress. This study sheds light on exploring and answering the molecular target of DA against target insects.

The blockade of the PD-1/PD-L1 immune checkpoint pathway with small molecules is an emerging immunotherapeutic approach. A novel series of 4-phenylindoline derivatives were synthesized, and their inhibitory activity against the PD-1/PD-L1 protein-protein interaction (PPI) was evaluated through a homogenous time-resolved fluorescence (HTRF) assay. Among them, A20 and A22 exhibited potent activity with IC50 values of 17 nM and 12 nM, respectively. Furthermore, A20 showed the promising inhibitory activity against the PD-1/PD-L1 interaction with the EC50 value of 0.43 μM in a co-culture model of PD-L1/TCR Activator-expressing CHO cells and PD-1-expressing Jurkat cells. Besides, the structure-activity relationships (SAR) of the novel synthesized 4-phenylindoline derivatives was concluded, and the binding mode of A22 with the PD-L1 dimer was analyzed by molecular simulation and docking, demonstrating that the N-atom in the side chain of indoline fragment could interact with the amino acid residue of the PD-L1 protein to lead to the potent inhibitory activity. This study provided a new insight for further drug design.

Aberrant activation of the endosomal Toll-like receptor 7 (TLR7) has been implicated in myriad autoimmune diseases and is an established therapeutic target in such conditions. Development of diverse TLR7 antagonists is mainly accomplished through random screening. To correlate human TLR7 (hTLR7) antagonistic activity with the structural features in different chemotypes, we derived a hypothetical binding model based on molecular docking analysis along with molecular dynamics (MD) simulations study. The binding hypothesis revealed different pockets, grooves and a central cavity where ligand-receptor interaction with specific residues through hydrophobic and hydrogen bond interactions take place, which correlate with TLR7 antagonistic activity thus paving the way for rational design using varied chemotypes. Based on the structural insight thus gained, TLR7 antagonists with quinazoline were designed to understand the effect of engagement of these pockets as well as boundaries of the chemical space associated with them. The newly synthesized most potent hTLR7 antagonist, i.e. compound 63, showed IC50 value of 1.03 ± 0.05 μM and was validated by performing primary assay in human plasmacytoid dendritic cells (pDC) (IC50pDC: 1.42 μM). The biological validation of the synthesized molecules was performed in TLR7-reporter HEK293 cells as well as in human plasmacytoid dendritic cells (pDCs). Our study provides a rational design approach thus facilitating further development of novel small molecule hTLR7 antagonists based on different chemical scaffolds.

Gliotoxin (GT) is a fungal secondary metabolite that has attracted great interest due to its high biological activity since it was discovered by the 1930s. It exhibits a unique structure that contains a N-C = O group as the characteristics of the classical PSII inhibitor. However, GT\'s phytotoxicity, herbicidal activity and primary action targets in plants remain hidden. Here, it is found that GT can cause brown or white leaf spot of various monocotyledonous and dicotyledonous plants, being regarded as a potential herbicidal agent. The multiple sites of GT action are located in two photosystems. GT decreases the rate of oxygen evolution of PSII with an I 50 value of 60 µM. Chlorophyll fluorescence data from Chlamydomonas reinhardtii cells and spinach thylakoids implicate that GT affects both PSII electron transport at the acceptor side and the reduction rate of PSI end electron acceptors\' pool. The major direct action target of GT is the plastoquinone QB-site of the D1 protein in PSII, where GT inserts in the QB binding niche by replacing native plastoquinone (PQ) and then interrupts electron flow beyond plastoquinone QA. This leads to severe inactivation of PSII RCs and a significant decrease of PSII overall photosynthetic activity. Based on the simulated modeling of GT docking to the D1 protein of spinach, it is proposed that GT binds to the-QB-site through two hydrogen bonds between GT and D1-Ser264 and D1-His252. A hydrogen bond is formed between the aromatic hydroxyl oxygen of GT and the residue Ser264 in the D1 protein. The 4-carbonyl group of GT provides another hydrogen bond to the residue D1-His252. So, it is concluded that GT is a novel natural PSII inhibitor. In the future, GT may have the potential for development into a bioherbicide or being utilized as a lead compound to design more new derivatives.

The skeletal muscle ryanodine receptor (RyR1) proteins are intracellular calcium (Ca2+) release channels on the membrane of the sarcoplasmic reticulum (SR) and required for skeletal muscle excitation-contraction coupling. Homer (Vesl) is a family of scaffolding proteins that modulate target proteins including RyRs (ryanodine receptors), mGluRs (group 1 metabotropic glutamate receptors) and IP3Rs (inositol-1,4,5-trisphosphate receptors) through a conserved EVH1 (Ena/VASP homology 1) domain. Here, we examined the interaction between Homer1 EVH1 domain and RyR1 by co-immunoprecipitation, continuous sucrose density-gradient centrifugation, and bio-layer interferometry binding assay at different Ca2+ concentrations. Our results show that there exists a high-affinity binding between the Homer1 EVH1 domain and RyR1, especially at 1 mM of Ca2+. Based on our data and the known structures of Homer1 EVH1 domain and RyR1, we found two consensus proline-rich sequences in the structure of RyR1, PPHHF and FLPPP, and proposed two corresponding binding models to show mechanisms of recognition different from those used by other proline-rich motifs. The side proline residues of two proline-rich motifs from RyR1 are away from the hydrophobic surface of Homer1 EVH1, rather than buried in this hydrophobic surface. Our results provide evidence that Homer1 regulates RyR1 by direct interaction.

Ligand binding dynamics and the concept of drug-target residence time are essential factors in the development of novel drugs. Conventional ligand binding assays, which usually collect end-point data, do not provide abundant information regarding the ligand binding kinetics. Therefore, novel methods that allow on-line monitoring of ligand binding processes have to be developed and implemented for drug discovery studies. In this study, we provide a short overview of novel possibilities to characterize ligand binding dynamics to different G protein-coupled receptors (GPCRs). Special attention has been paid to the ligand binding to melanocortin 4 receptors and to the development of a fluorescence anisotropy-based assay system using receptors in budded baculovirus particles. It has been shown that ligand binding to melanocortin 4 receptors occurs to tandemly arranged interconnecting ligand binding sites and that the conventional equilibrium usually cannot be achieved in this system. Therefore, the apparent potencies of the same ligand may differ by up to four orders of magnitude, depending on the experimental conditions and the reporter ligand used.