A method for rapid screening of interactions of pharmacologically active compounds with albumin

Majcher, Aldona; Lewandrowska, Anna; Herold, Franciszek; Stefanowicz, Jacek; Słowiński, Tomasz; Mazurek, Aleksander P.; Wieczorek, Stefan A.; Hołyst, Robert

doi:10.1016/j.aca.2014.12.008

Cited by 10 publications

(7 citation statements)

References 36 publications

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“…However, in case of observation of molecular motion, careful choice of probes is crucial: the fact that charge screening is sufficient to elasticize the polyelectrolyte chains does not preclude attractive interaction between probe and crowder of opposite charges. In fact, comparison of the diffusion rates predicted by the model given hereby with diffusion coefficients measured for probes interacting with the crowders (of opposite charges) should allow for quantitative elucidation of the equilibrium and rate constants of probe attachment to the crowders [ 68 , 69 ]. This opens a way towards systematic studies of the influence of electrostatic attraction on (macro)molecular mobility and reactivity in crowded, biologically relevant systems.…”

Section: Discussionmentioning

confidence: 99%

Motion of Molecular Probes and Viscosity Scaling in Polyelectrolyte Solutions at Physiological Ionic Strength

et al. 2016

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We investigate transport properties of model polyelectrolyte systems at physiological ionic strength (0.154 M). Covering a broad range of flow length scales—from diffusion of molecular probes to macroscopic viscous flow—we establish a single, continuous function describing the scale dependent viscosity of high-salt polyelectrolyte solutions. The data are consistent with the model developed previously for electrically neutral polymers in a good solvent. The presented approach merges the power-law scaling concepts of de Gennes with the idea of exponential length scale dependence of effective viscosity in complex liquids. The result is a simple and applicable description of transport properties of high-salt polyelectrolyte solutions at all length scales, valid for motion of single molecules as well as macroscopic flow of the complex liquid.

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Section: Discussionmentioning

confidence: 99%

Motion of Molecular Probes and Viscosity Scaling in Polyelectrolyte Solutions at Physiological Ionic Strength

et al. 2016

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“…The equilibrium constant can be determined by an independent technique, e.g., TDA. [34][35][36] The rest of the parameters (t A , t B , t D , t AE and b) whose values depend on the structure of the focal volume and the micellar concentration in formula (8) can be exactly calculated and then fixed during the fitting procedure. The target quantity R is the only free model parameter in G a (t) to fit, besides the triplet-state parameters p and t t .…”

Section: Approximate Form Of the Autocorrelation Functionmentioning

confidence: 99%

“…To test this method, we also determine the equilibrium constant by Taylor dispersion analysis (TDA). [34][35][36] Then, we use FCS to investigate the kinetics of complex formation, i.e., the association and dissociation rates. It is feasible if the chemical relaxation rate (definition in the Materials and methods section) is reduced below the limit of fast reaction, which can be usually achieved in the case of diluted micellar solutions because the relaxation rate depends linearly on the concentration of micelles.…”

Section: Introductionmentioning

confidence: 99%

Determination of equilibrium and rate constants for complex formation by fluorescence correlation spectroscopy supplemented by dynamic light scattering and Taylor dispersion analysis

et al. 2016

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The equilibrium and rate constants of molecular complex formation are of great interest both in the field of chemistry and biology. Here, we use fluorescence correlation spectroscopy (FCS), supplemented by dynamic light scattering (DLS) and Taylor dispersion analysis (TDA), to study the complex formation in model systems of dye-micelle interactions. In our case, dyes rhodamine 110 and ATTO-488 interact with three differently charged surfactant micelles: octaethylene glycol monododecyl ether CE (neutral), cetyltrimethylammonium chloride CTAC (positive) and sodium dodecyl sulfate SDS (negative). To determine the rate constants for the dye-micelle complex formation we fit the experimental data obtained by FCS with a new form of the autocorrelation function, derived in the accompanying paper. Our results show that the association rate constants for the model systems are roughly two orders of magnitude smaller than those in the case of the diffusion-controlled limit. Because the complex stability is determined by the dissociation rate constant, a two-step reaction mechanism, including the diffusion-controlled and reaction-controlled rates, is used to explain the dye-micelle interaction. In the limit of fast reaction, we apply FCS to determine the equilibrium constant from the effective diffusion coefficient of the fluorescent components. Depending on the value of the equilibrium constant, we distinguish three types of interaction in the studied systems: weak, intermediate and strong. The values of the equilibrium constant obtained from the FCS and TDA experiments are very close to each other, which supports the theoretical model used to interpret the FCS data.

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“…Although noncovalent interactions are generally much weaker than covalent bonding, they significantly affect the physicochemical properties of matters and play a crucial role in the processes of complex formation in biochemical and supramolecular systems . Determination of equilibrium and rate constants of these interactions can provide quantitative information on these systems. , Complex formation processes are often extremely fast (up to the diffusion-controlled limit) and require highly advanced techniques to study them quantitatively. − The main methods used previously, e.g., NMR, titration, sometime gave relatively inconsistent results since individuals’ behaviors of molecules were averaged in the experiments. ,, Single-molecule techniques are ideal tools for the characterization of molecular interactions. One of the most promising experimental solutions is the fluorescence correlation spectroscopy (FCS). ,− In FCS, fluctuations of fluorescence originating from probes diffusing through an observation volume are recorded.…”

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confidence: 99%

Nanoscopic Approach to Quantification of Equilibrium and Rate Constants of Complex Formation at Single-Molecule Level

Zhang

Sisamakis

Sozański

et al. 2017

J. Phys. Chem. Lett.

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Equilibrium and rate constants are key descriptors of complex-formation processes in a variety of chemical and biological reactions. However, these parameters are difficult to quantify, especially in the locally confined, heterogeneous, and dynamically changing living matter. Herein, we address this challenge by combining stimulated emission depletion (STED) nanoscopy with fluorescence correlation spectroscopy (FCS). STED reduces the length-scale of observation to tens of nanometres (2D)/attoliters (3D) and the time-scale to microseconds, with direct, gradual control. This allows one to distinguish diffusional and binding processes of complex-formation, even at reaction rates higher by an order of magnitude than in confocal FCS. We provide analytical autocorrelation formulas for probes undergoing diffusion-reaction processes under STED condition. We support the theoretical analysis of experimental STED-FCS data on a model system of dye-micelle, where we retrieve the equilibrium and rates constants. Our work paves a promising way toward quantitative characterization of molecular interactions in vivo.

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A method for rapid screening of interactions of pharmacologically active compounds with albumin

Cited by 10 publications

References 36 publications

Motion of Molecular Probes and Viscosity Scaling in Polyelectrolyte Solutions at Physiological Ionic Strength

Motion of Molecular Probes and Viscosity Scaling in Polyelectrolyte Solutions at Physiological Ionic Strength

Determination of equilibrium and rate constants for complex formation by fluorescence correlation spectroscopy supplemented by dynamic light scattering and Taylor dispersion analysis

Nanoscopic Approach to Quantification of Equilibrium and Rate Constants of Complex Formation at Single-Molecule Level

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