Probing Protein–Ligand Methyl−π Interaction Geometries through Chemical Shift Measurements of Selectively Labeled Methyl Groups

Fragment-based drug design is heavily dependent on the
optimization
of initial low-affinity binders. Herein we introduce an approach that
uses selective labeling of methyl groups in leucine and isoleucine
side chains to directly probe methyl−π contacts, one
of the most prominent forms of interaction between proteins and small
molecules. Using simple NMR chemical shift perturbation experiments
with selected BRD4-BD1 binders, we find good agreement with a commonly
used model of the ring-current effect as well as the overall interaction
geometries extracted from the Protein Data Bank. By combining both
interaction geometries and chemical shift calculations as fit quality
criteria, we can position dummy aromatic rings into an AlphaFold model
of the protein of interest. The proposed method can therefore provide
medicinal chemists with important information about binding geometries
of small molecules in fast and iterative matter, even in the absence
of high-resolution experimental structures.