Ubiquitin, Cavitand, and NMR: 15N-1H HSQC Analysis of Cavitand Binding Domains on Ubiquitin

Ubiquitin and its derivatives through side chain modification are present in many of the degradation pathways for proteins found in neurodegenerative diseases. A non-toxic method for determining the amount of these proteins could have biosensing applications, but the binding domains of such proteins are not well characterized. 15N-1H HSQC analysis of ubiquitin’s interactions with different concentrations of a small negatively charged host molecule called cavitand provides insight into the binding interactions with side chains on the protein such as lysine and arginine. This is done comparing the HSQC spectra of different concentrations of cavitand and ubiquitin to see where the cavitand binds and what conformational changes are induced. The cavitand was found to bind to positively charged Lysine residues on the ubiquitin as well as non-polar side chains. Further conformational changes suggest binding in other areas, but further research is needed to elucidate these findings. Altering the qualities and concentration of the cavitand will provide insight into how these interactions occur. Once these interactions can be characterized and optimized, cavitand has good potential to be a biosensor for post translational modifications of ubiquitin.