Heparin and Heparan Sulfate Binding of the Antiparasitic Drug Imidocarb: Circular Dichroism Spectroscopy, Isothermal Titration Calorimetry, and Computational Studies

Abstract

This study is aimed to assess the binding interaction between the antiparasitic cationic drug imidocarb (IMD) and sulfated glycosaminoglycans (GAGs), the ubiquitious nonprotein macromolecules of living organisms. These complex, heterogeneous polyanions are the integral constituents of cell membranes and the extracellular matrix and display affinity toward basic compounds, the binding of which may affect their biological functions. Exciton-type circular dichroism (CD) spectroscopic features measured at low salt concentration verify the heparin and heparan sulfate binding of IMD, which occurs in a cooperative manner by association of several drug molecules to a disaccharide unit. Isothermal titration calorimetry (ITC) measurements reassured the heparin interaction, resulting in a K value in the low micromolar range. In contrast, when considering high molar excess of the heparin-binding sites, closer resembling in vivo conditions, an entirely different CD signature was induced, suggesting a shift from the oligo- to monomeric binding mode. This observation was also supported by ITC measurements using an identical sample setup. To better mimic in vivo conditions, several measurements were performed in physiological salt concentration ranges. On the basis of these, the inter- and intramolecular origin of CD activity observed under low- and high-salt conditions refer to electrostatically held oligomeric and intermolecular H-bonded monomeric drug-GAG adducts, respectively. To complement the experimental data, quantum chemical calculations were performed to assess the photophysical and conformational properties of IMD, indicating the existence of nonlinear, nonplanar interconverting conformer populations. Such a structural flexibility may be important in the multiple, cooperative binding of IMD to sterically adjacent GAG sites.

Mind Your Dye: The Amyloid Sensor Thioflavin T Interacts with Sulfated Glycosaminoglycans Used To Induce Cross-β-Sheet Motifs

“…Simultaneously with the blue shift of the absorption peak, a negative–positive band pair was developed in the CD spectrum, showing a minimum at 404 and a maximum at 384 nm (Figure ). Its biphasic pattern is highly characteristic of the chiral intermolecular exciton coupling mechanism, which occurs through the space between two (or more) excitation dipole moments held in a chiral orientation relative to each other. − The sign of the resulting exciton peaks is determined by the screw sense (handedness) between the interacting transition moments: counterclockwise (left-handed) orientation leads to a longer-wavelength negative and a shorter-wavelength positive exciton band pair and vice versa. , The exciton components are displayed in the region of the respective absorption band and the zero intercept point between them concurs with the absorption maximum. Taking these features into consideration, the heparin-induced CD spectrum of ThT stems from the coupled oscillator interaction between the card-pack stacks of the dye molecules attached regularly to the helically arranged anionic sites of the polymer host.…”

NMR Characterization of the Interactions Between Glycosaminoglycans and Proteins

“…The interactions between GAGs and proteins are closely related to many factors, including saccharide unit composition, degree of sulfation, sulfation pattern, chain length, monosaccharide ring conformation and glycosidic linkage. The research methods used to characterize the interaction between GAGs and proteins mainly include gel electrophoresis (GE) (Nogueira et al, 2019), affinity chromatography (AC) (Sandoval et al, 2020), surface plasmon resonance (SPR) (Przybylski et al, 2020), biological layer interferometry (BLI) (Xiao et al, 2016), isothermal titration (ITC) (Zsila et al, 2018), microarray methods (Pomin and Wang, 2018b), crystal diffraction methods (X-ray) (Dahms et al, 2015), mass spectrometry (MS) (Yang and Chi, 2017), and nuclear magnetic resonance spectroscopy (NMR) (Kato and Peters, 2017). NMR is an insensitive technique compared with other analytical method for the study of interactions between biomolecules.…”

Understanding the Interaction of Polyelectrolyte Architectures with Proteins and Biosystems

“…Thus, linear plots of log K b versus log c s are found in a number of investigations [65] . The number of quantitative studies employing Equation (3), however, is rather small given the obvious importance of GAGs as biomaterials [19, 155, 156] . It is important to note that electrostatic interactions with heparin are already used in medical applications.…”