Solid-supported proteins in the liquid chromatography domain to probe ligand-target interactions
- 1Instituto de Química, Departamento de Química Orgânica, Universidade Federal Fluminense, Brazil
- 2Faculty of Philosophy, Sciences and Languages ??of Ribeirão Preto, University of São Paulo, Brazil
- 3Departamento de Química, Universidade Federal de São Carlos, Brazil
Ligand-target interactions play a central role in drug discovery processes because these interactions are crucial in biological systems. Small molecules-proteins interactions can regulate and modulate protein function and activity through conformational changes. Therefore, bioanalytical tools to screen new ligands have focused mainly on probing ligand-target interactions. These interactions have been evaluated by using solid-supported proteins, which provide advantages like increased protein stability and easier protein extraction from the reaction medium, which enables protein reuse. In some specific approaches, precisely in the ligand fishing assay, the bioanalytical method allows the ligands to be directly isolated from complex mixtures, including combinatorial libraries and natural products extracts without prior purification or fractionation steps. Most of these screening assays are based on liquid chromatography separation, and the binding events can be monitored through on-line or off-line methods. In the on-line approaches, solid supports containing the immobilized biological target are used as chromatographic columns most of the time. Several terms have been used to refer to such approaches, such as weak affinity chromatography, high-performance affinity chromatography, on-flow activity assays, and high-performance liquid affinity chromatography. On the other hand, in the off-line approaches, the binding event occurs outside the liquid chromatography system and may encompass affinity and activity-based assays in which the biological target is immobilized on magnetic particles or monolithic silica, among others. After the incubation step, the supernatant or the eluate from the binding assay is analyzed by liquid chromatography coupled to various detectors. Regardless of the selected bioanalytical approach, the use of solid supported proteins has significantly contributed to the development of automated and reliable screening methods that enable ligands to be isolated and characterized in complex matrixes without purification, thereby reducing costs and avoiding time-laborious steps. This review provides a critical overview of recently developed assays.
Keywords: Bioaffinity chromatography, Ligand screening, Ligand-target interactions, zonal bioaffinity chromatography, frontal bioaffinity chromatography, Ligand fishing
Received: 10 Sep 2019;
Accepted: 21 Oct 2019.
Copyright: © 2019 De Moraes, Cardoso and Cass. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Prof. Marcela C. De Moraes, Instituto de Química, Departamento de Química Orgânica, Universidade Federal Fluminense, Niterói, Brazil, firstname.lastname@example.org