Deciphering the molecular details of interactions between anti-COVID drugs and functional human proteins: in silico approach.

The molecular docking calculations have been employed to investigate the interactions a set of proteins with the repurposed anti-COVID drugs. The position of the therapeutic agents within the protein structure was dependent on a particular drug-protein system and varied from the binding cleft to the periphery of the polypeptide chain. Interactions involved in the drug-protein complexation includes predominantly hydrogen bonding and hydrophobic contacts.

Fӧrster resonance energy transfer analysis of amyloid state of proteins.

The Förster resonance energy transfer (FRET) is a well-established and versatile spectroscopic technique extensively used for exploring a variety of biomolecular interactions and processes. The present review is intended to cover the main results of our FRET studies focused on amyloid fibrils, a particular type of disease-associated protein aggregates. Based on the examples of several fibril-forming proteins including insulin, lysozyme and amyloidogenic variants of N-terminal fragment of apolipoprotein A-I, it was demonstrated that: (i) the two- and three-step FRET with the classical amyloid marker Thioflavin T as an input donor has a high amyloid-sensing potential and can be used to refine the amyloid detection assays; (ii) the intermolecular time-resolved and single-molecule pulse interleaved excitation FRET can give quantitative information on the nucleation of amyloid fibrils; (iii) FRET between the membrane fluorescent probes and protein-associated intrinsic or extrinsic fluorophores is suitable for monitoring the membrane binding of fibrillar proteins, exploring their location relative to lipid-water interface and restructuring on a lipid matrix; (iv) the FRET-based distance estimation between fibril-bound donor and acceptor fluorophores can serve as one of the verification criteria upon structural modeling of amyloid fibrils.

Three-step Förster resonance energy transfer on an amyloid fibril scaffold.

The present study provides evidence that the energy transfer chain consisting of the benzothiazole dye Thioflavin T as an input donor, a phosphonium dye TDV and a squaraine dye SQ4 as mediators, and one of the three squaraines SQ1/2/3 as an output acceptor displays an excellent amyloid-sensing ability when applied to differentiating between the amyloid and non-fibrillized states of insulin. The ensemble of fluorophores offers the advantages of a large effective Stokes shift (∼240 nm), well-resolved 3D fluorescence patterns and strong enhancement of the terminal fluorescence (up to two orders of magnitude). The occurrence of multistep energy transfer on an amyloid fibril scaffold opens new possibilities for the more sensitive detection of fibrillar protein assemblies and their applications in nanophotonics.

Probing the interactions of novel europium coordination complexes with serum albumin.

Molecular interactions between novel europium coordination complexes (EC) possessing superior cytotoxic activity and bovine serum albumin (BSA), the most prominent representative of plasma proteins, were assessed using fluorescence spectroscopy and molecular docking techniques. Cumulative results from fluorescent probe binding, fluorescence quenching and Förster resonance energy transfer studies revealed that the europium complexes V4 and V8 do not perturb the BSA structure, while V3, V5, and V7 induce partial unfolding of the polypeptide chain. Molecular docking studies coupled with analysis of the three-dimensional structure of the BSA-EC complexes showed that V4 and V8 reside in the vicinity of the protein IIA subdomain (Sudlow's site I), while V3, V5 and V5 were localized predominantly in the BSA IIIA subdomain (Sudlow's site II).

Aggregation behavior of novel heptamethine cyanine dyes upon their binding to native and fibrillar lysozyme.

Two newly synthesized symmetrical heptamethine cyanine dyes, AK7-5 and AK7-6, absorbing in the region of low autofluorescence of biological samples, have been tested for their ability to detect proteins aggregated into amyloid fibrils. In aqueous solution these probes possess three absorption bands corresponding to the monomer, dimer and H-aggregate species. The association of the dye with fibrillar lysozyme was followed by the enhancement of the monomer band and the reduction of the H-band.

Fluorescence monitoring of the effect of oxidized lipids on the process of protein fibrillization.

The kinetics of lysozyme and insulin amyloid formation in the presence of the oxidized phospholipids (oxPLs) was investigated using Thioflavin T fluorescence assay. The kinetic parameters of fibrillization process (lag time and apparent rate constant) have been determined upon varying the following experimental parameters: the type of lipid assemblies (premicellar aggregates and lipid bilayer vesicles), pH, temperature and lipid-to-protein molar ratio. It was found that oxPLs premicellar aggregates induced the more pronounced increase of the maximum Thioflavin T fluorescence, which is proportional to the extent of fibril formation, compared to the vesicles composed of the oxidized and unoxidized lipids.

Novel benzanthrone probes for membrane and protein studies.

The applicability of a series of novel benzanthrone dyes to monitoring the changes in physicochemical properties of lipid bilayer and to differentiating between the native and aggregated protein states has been evaluated. Based on the quantitative parameters of the dye-membrane and dye-protein binding derived from the fluorimetric titration data, the most prospective membrane probes and amyloid tracers have been selected from the group of examined compounds. Analysis of the red edge excitation shifts of the membrane- and amyloid-bound dyes provided information on the properties of benzanthrone binding sites within the lipid and protein matrixes.

Fluorescence Investigation of Interactions Between Novel Benzanthrone Dyes and Lysozyme Amyloid Fibrils.

A series of novel fluorescent benzanthrone dyes have been tested for their ability to identify and characterize fibrillar aggregates of lysozyme prepared by protein denaturation in concentrated ethanol solution (F(eth)) or acidic buffer (F(ac)). Quantitative parameters of the dye association with native and fibrillar protein have been derived from the results of fluorimetric titration. The binding characteristics proved to be different for F(eth)- and F(ac)-bound benzanthrones, highlighting the dye sensitivity to the distinctions in fibril morphology.