Protein−protein interaction is the fundamental step of biological signal transduction. Interacting proteins find each other by diffusion. To gain insight into diffusion under the crowded conditions found in cells, researchers in the Pielak Group used nuclear magnetic resonance spectroscopy (NMR) to measure the effects of solvent additives on the translational and rotational diffusion of the 7.4 kDa globular protein, chymotrypsin inhibitor 2.
The additives were glycerol and the macromolecular crowding agent, polyvinyl pyrrolidone (PVP). As published in the Journal of Physical Chemistry B, both translational diffusion and rotational diffusion decrease with increasing solution viscosity. For glycerol, the decrease obeys the Stokes−Einstein and Stokes−Einstein Debye laws. Three types of deviation are observed for PVP: the decrease in diffusion with increased viscosity is less than predicted, this negative deviation is greater for rotational diffusion, and the negative deviation increases with increasing PVP molecular weight.
Fibrils of the intrinsically disordered protein α-synuclein are hallmarks of Parkinson’s disease. The fluorescent dye thioflavin T is often used to characterize fibrillation, but this assay may not provide quantitative information about structure and mechanism. To gain such information, researchers in the Pielak Group, as reported in the journal Biochemistry, incorporated the 19F-labeled amino acid, 3-fluorotyrosine, into recombinant human α-synuclein at its endogenous tyrosine residues. 19F nuclear magnetic resonance spectroscopy was used to study these labeled α-synucleins.
Although dye binding and 19F NMR data show that 1 mM SDS and 1 mM spermine accelerate aggregation compared to buffer alone, only the NMR data indicate that the species formed in SDS are smaller than those formed in buffer or buffer plus spermine. Thus, 19F NMR spectroscopy can provide quantitative, residue-level, information about protein structure, binding, and fibrillation.