![]() ![]() In a broader biological context, these associations are linked and interconnected in complex networks that allow sensitive and precise developmental programs and responses to environmental cues, and that are altered in disease states. ![]() Their thermodynamics provides information critical for deriving a fundamental understanding of molecular functions. Molecular associations lie at the heart of biology. We apply this framework and explain underlying fundamental concepts through experimental examples with the RNA-binding protein Puf4. 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. Moreover, several reported affinities could be concluded to be incorrect, thereby impacting biological interpretations. 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. 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. Quantitative measurements of biomolecule associations are central to biological understanding and are needed to build and test predictive and mechanistic models. ![]()
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