Utilizing Molecular Probes to Examine Protein Structure-Function Relationships

Within the cell, the majority of functional roles are carried out by polypeptide macromolecules commonly referred to as proteins. Over the past 100 years, various methodologies to examine the high-resolution structural characteristics of proteins have been developed and utilized, including x-ray crystallography, nuclear magnetic resonance spectroscopy, and cryogenic electron microscopy. The structural information obtained has yielded much insight into the mechanisms by which many of these proteins function; however, limitations to the nature of the structural data have provided challenges when attempting to elucidate protein structure-function relationships and dynamics. Owing to its nature, x-ray crystallography provides what can be described as a “snapshot” of the most favorable protein conformation or conformations in a particular crystal, which is affected by both the components of the solution as well as the crystallographic contacts that are necessary for the formation of the lattice. In this work, three different proteins have been examined crystallographically and their structures perturbed by the addition of molecular probes ranging in size from the miniscule molecular oxygen (hemoglobin), through the small molecule range of 50-1000 daltons (MDM2 RING domain), and well into the macromolecular range of \u3e1 kilodalton (PU.1 transcription factor DNA-binding domain). Variances observed in the determined x-ray structures yielded valuable insight into the mechanisms by which these proteins function, as well as a deeper understanding of the methods by which they may be targeted for future treatment of human disease