Journal Article

How to measure and evaluate binding affinities

AlSadhan, Ishraq, Jarmoskaite, Inga, Herschlag, Daniel, Vaidyanathan, Pavanapuresan P

2020

Overview

Quantitative measurements of biomolecule associations are central to biological understanding and are needed to build and test predictive and mechanistic models. Given the advances in high-throughput technologies and the projected increase in the availability of binding data, we found it especially timely to evaluate the current standards for performing and reporting binding measurements. A review of 100 studies revealed that in most cases essential controls for establishing the appropriate incubation time and concentration regime were not documented, making it impossible to determine measurement reliability. Moreover, several reported affinities could be concluded to be incorrect, thereby impacting biological interpretations. Given these challenges, we provide a framework for a broad range of researchers to evaluate, teach about, perform, and clearly document high-quality equilibrium binding measurements. We apply this framework and explain underlying fundamental concepts through experimental examples with the RNA-binding protein Puf4.

• Publisher: eLife Science Publications, Ltd,eLife Sciences Publications Ltd,eLife Sciences Publications, Ltd
• Subject: Analysis / binding affinity / Biochemistry and Chemical Biology / Biology / Case-Control Studies / dissociation constant / Earthquakes / Experiments / Kinetics / Ligands / Measurement / Protein Binding / protein‐ligand interaction / Reproducibility of Results / Ribonucleic acid / RNA / RNA binding protein / RNA-Binding Proteins - metabolism / RNA-protein interactions / Saccharomyces cerevisiae Proteins - metabolism / Thermodynamics / Tools and Resources / Values