Native Structure-Based Peptides as Potential Protein-Protein Interaction Inhibitors of SARS-CoV-2 Spike Protein and Human ACE2 Receptor.

Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) is a positive-strand RNA virus that causes severe respiratory syndrome in humans, which is now referred to as coronavirus disease 2019 (COVID-19). Since December 2019, the new pathogen has rapidly spread globally, with over 65 million cases reported to the beginning of December 2020, including over 1.5 million deaths. Unfortunately, currently, there is no specific and effective treatment for COVID-19. As SARS-CoV-2 relies on its spike proteins (S) to bind to a host cell-surface receptor angiotensin-converting enzyme-2(ACE2), and this interaction is proved to be responsible for entering a virus into host cells, it makes an ideal target for antiviral drug development. In this work, we design three very short peptides based on the ACE2 sequence/structure fragments, which may effectively bind to the receptor-binding domain (RBD) of S protein and may, in turn, disrupt the important virus-host protein-protein interactions, blocking early steps of SARS-CoV-2 infection. Two of our peptides bind to virus protein with affinity in nanomolar range, and as very short peptides have great potential for drug development.

Thermodynamic contribution of iodine atom to the binding of heterogeneously polyhalogenated benzotriazoles by the catalytic subunit of human protein kinase CK2.

A series of novel benzotriazole derivatives containing iodine atom(s) were synthesized. The binding of these compounds to the catalytic subunit of human protein kinase CK2 was evaluated using differential scanning fluorimetry. The obtained thermodynamic data were compared with those determined previously for the brominated and chlorinated benzotriazole analogues to get a deeper insight into the thermodynamic contribution of iodine substitution to the free energy of ligand binding. We have shown that iodine atom(s) attached to the benzene ring of benzotriazole enhance(s) its binding by the target protein. This effect is the strongest when two iodine atoms are attached at positions peripheral to the triazole ring, which according to the structures deposited in protein data bank may be indicative for the formation of the halogen bond between iodine and carbonyl groups of residues located in the hinge region of the protein. Finally, quantitative structure-activity relationship analysis pointed the solute hydrophobicity as the main factor contributing to the binding affinity.

The halogenation of natural flavonoids, baicalein and chrysin, enhances their affinity to human protein kinase CK2.

A series of halogenated derivatives of natural flavonoids: baicalein and chrysin were designed and investigated as possible ligands for the catalytic subunit of tumor-associated human kinase CK2. Thermal shift assay method, in silico modeling, and high-performance liquid chromatography-derived hydrophobicity together with IC50 values determined in biochemical assay were used to explain the ligand affinity to the catalytic subunit of human protein kinase CK2. Obtained results revealed that substitution of baicalein and chrysin with halogen atom increases their binding affinity to hCK2α, and for 8-chlorochrysin the observed effect is even stronger than for the reference CK2 inhibitor-4,5,6,7-tetrabromo-1H-benzotriazole. The cytotoxic activities of the baicalein and chrysin derivatives in the in vitro model have been evaluated for MV4-11 (human biphenotypic B myelomonocytic leukemia), A549 (human lung adenocarcinoma), LoVo (human colon cancer), and MCF-7 (human breast cancer) as well as on the nontumorigenic human breast epithelial MCF-10A cell lines. Among the baicalein derivatives, the strongest cytotoxic effect was observed for 8-bromobaicalein, which exhibited the highest activity against breast cancer cell line MCF-7 (IC50 10 ± 3 µM). In the chrysin series, the strongest cytotoxic effect was observed for unsubstituted chrysin, which exhibited the highest activity against leukemic cell line MV4-11 (IC50 10 ± 4 µM).

A competition between hydrophobic and electrostatic interactions in protein-ligand systems. Binding of heterogeneously halogenated benzotriazoles by the catalytic subunit of human protein kinase CK2.

A series of chlorine-substituted benzotriazole derivatives, representing all possible substitution patterns of halogen atoms attached to the benzotriazole benzene ring, were synthetized as potential inhibitors of human protein kinase CK2. Basic ADME parameters for the free solutes (hydrophobicity, electronic properties) together with their binding affinity to the catalytic subunit of protein kinase CK2 were determined with reverse-phase HPLC, spectrophotometric titration, and Thermal Shift Assay Method, respectively. The analysis of position-dependent thermodynamic contribution of a chlorine atom attached to the benzotriazole ring confirmed the previous observation for brominated benzotriazoles, in which substitution at positions 5 and 6 with bromine was found crucial for ligand binding. In all tested halogenated benzotriazoles the replacement of Br with Cl decreases the hydrophobicity, while the electronic properties remain virtually unaffected. Supramolecular architecture identified in the just resolved crystal structures of three of the four possible dichloro-benzotriazoles shows how substitution distant from the triazole ring affects the pattern of intermolecular interactions. Summarizing, the benzotriazole benzene ring substitution pattern has been identified as the main driver of ligand binding, predominating the non-specific hydrophobic effect.

Urinary tract infection during pregnancy affects the level of leptin, ghrelin and insulin in maternal and placental blood.

AIM: We examined ghrelin, leptin and insulin in maternal blood during normal pregnancy and pregnancy complicated by urinary tract infection (UTI), as well as in cord blood at labor. METHODS: A total of 36 delivering women with history of UTI during the third trimester of pregnancy were enrolled in the study; 12 healthy pregnant women served as a control. Infection markers (CRP and procalcitonin) were determined in maternal blood during the course of UTI and at labor. Ghrelin, leptin and insulin were determined during labor in venous maternal and in umbilical cord blood. RESULTS: We found negative correlation between infection markers in maternal blood during UTI, and level of tested hormones in cord blood, indicating potential risk of placental impairment due to energetic imbalance. We noted lower level of leptin in mothers with UTI and no change in leptin from umbilical blood comparing subjects with and without UTI. Low level of ghrelin was observed in maternal and cord blood when pregnancy was complicated by UTI. Insulin concentrations were high in mothers with UTI and low in their newborn's cord blood. Increased maternal insulin level could indicate peripheral insulin resistance caused by the infection. CONCLUSION: UTI during pregnancy affects the concentration of hormones responsible for regulating energetic homeostasis within the placenta.

Effect of histidine protonation state on ligand binding at the ATP-binding site of human protein kinase CK2.

Histidine residues contribute to numerous molecular interactions, owing to their structure with the ionizable aromatic side chain with pKa close to the physiological pH. Herein, we studied how the two histidine residues, His115 and His160 of the catalytic subunit of human protein kinase CK2, affect the binding of the halogenated heterocyclic ligands at the ATP-binding site. Thermodynamic studies on the interaction between five variants of hCK2α (WT protein and four histidine mutants) and three ionizable bromo-benzotriazoles and their conditionally non-ionizable benzimidazole counterparts were performed with nanoDSF, MST, and ITC. The results allowed us to identify the contribution of interactions involving the particular histidine residues to ligand binding. We showed that despite the well-documented hydrogen bonding/salt bridge formation dragging the anionic ligands towards Lys68, the protonated His160 also contributes to the binding of such ligands by long-range electrostatic interactions. Simultaneously, His 115 indirectly affects ligand binding, placing the hinge region in open/closed conformations.

Coronary Artery Disease and Inflammatory Activation Interfere with Peripheral Tissue Electrical Impedance Spectroscopy Characteristics-Initial Report.

BACKGROUND: The electrical properties of cells and tissues in relation to energy exposure have been investigated, presenting their resistance and capacitance characteristics. The dielectric response to radiofrequency fields exhibits polarization heterogeneity under pathological conditions. The aim of the study was to analyze the differences in changes in resistance and capacitance measurements in the range from 1 kHz to 1 MHz, combined with an assessment of the correlation between the results of electrical impedance spectroscopy (EIS) and inflammatory activation. METHODS: In the prospective study, EIS was performed on the non-dominant arm in 29 male patients (median (Q1-Q3) age of 69 (65-72)) with complex coronary artery disease and 10 male patients (median (Q1-Q3) age of 66 (62-69)) of the control group. Blood samples were collected for inflammatory index analysis. RESULTS: The logistic regression analysis revealed a negative correlation with inflammatory indexes, including neutrophil to lymphocyte ratio (NLR) in the CAD group in the frequency of 30 kHz (p = 0.038, r = -0.317) regarding EIS resistance measurements and a positive correlation in CAD group in the frequency of 10 kHz (p = 0.029, r = -0.354) regarding EIS capacitance. CONCLUSIONS: The bioelectric characteristics of peripheral tissues measured by resistance and capacitance in EIS differ in patients with coronary artery disease and in the control group. Electrical impedance spectroscopy reveals a statistically significant correlation with inflammatory markers in patients with CAD.

Efficacy evaluation of electric field frequency and temperature on dielectric properties of collagen cross-linked by glutaraldehyde.

Solid-state dielectric properties are reported for unmodified collagen (Col) and glutaraldehyde-modified collagen (Col-GA) over the frequency range from 100Hz to 100kHz and at temperatures from 25 to 145°C. In the full temperature and frequency range the average values of the relative permittivity and dielectric loss for Col samples are higher than those recorded for Col-GA samples. The peak temperature of these both parameters associated with the release of loosely bound water is around 73 and 77°C for Col and Col-GA samples, respectively. The activation energy for the reorientation and breaking of hydrogen bonds takes the values 32kJmol-1 for Col and 23kJmol-1 for Col-GA. The relative permittivity decrement and conductivity increment of Col-GA samples fall by 40 and 30% on average in the temperature range 25-75°C, as compared to Col samples. Dielectric properties of Col-GA may be helpful in designing scaffolds for tissue engineering.

Pore structure and dielectric behaviour of the 3D collagen-DAC scaffolds designed for nerve tissue repair.

The design and selection of a suitable scaffold with well-defined pores size distribution and dielectric properties are critical features for neural tissue engineering. In this study we use mercury porosimetry and the dielectric spectroscopy in the alpha-dispersion region of the electric field to determine the microarchitecture and activation energy of collagen (Col) modified by 2,3 dialdehyde cellulose (DAC). The scaffold was synthesized in three steps: (i) preparation of DAC by oxidation of cellulose, (ii) construction of a 3D Col sponge-shape or film, (iii) cross-linkage of the Col samples using DAC. The activation energy needed to break the bonds formed by water in the Col-DAC composite is approximately 2 times lower than that in the unmodified Col. In addition, the magnitude of conductivity for modified Col at 70°C is approximately 40% lower than that recorded for the unmodified Col. The largest fraction, of which at least 70% of the total pore volume comprises the sponge, is occupied by pores ranging from 20 to 100µm in size. The knowledge on the dielectric behaviour and microstructure of the Col-DAC scaffold may prove relevant to neural tissue engineering focused on the regeneration of the nervous system.

In vitro evaluation of the effect of tobacco smoke on rat cornea function.

The influence of tobacco smoke on the dielectric properties of rat cornea were measured in vitro over the frequency range of the electric field of 500Hz-100kHz and in temperatures of the air from 25 to 150°C. The temperature dependencies of the loss tangent for both healthy and smoky cornea represent the relation between the energy lost and the energy stored in the epithelium-stromal-endothelium systems of the cornea. The differences between the healthy and the smoky cornea concerned the temperature ranges in which there appeared the decomposition of loosely-bound water and ß-relaxation associated with polar side-chains relaxations on protein molecules of this tissue. The effect of smoke is manifested as a shift of the loss tangent peaks of these two processes towards higher temperatures, when compared with the control. The results are interpreted as caused by the toxic compounds of the tobacco smoke leading to higher ion transport in the nonhomogeneous structure of the cornea when compared to that of the control tissue. The activation energy of conductivity were similar for the healthy and smoky cornea as a consequence of the braking of hydrogen and Van der Waals bonds between loosely bound water, and the proteins of channels in the epithelium and endothelium. Recognition of the effect of frequency and temperature on the dielectric behaviour of the smoky cornea may be of interest for disease characterization of this tissue.