Protein nuclear magnetic resonance under physiological conditions

Abstract

Almost everything we know about protein biophysics comes from studies on purified proteins in dilute solution. Most proteins, however, operate inside cells where the concentration of macromolecules can be >300 mg/mL. Although reductionism-based approaches have served protein science well for more than a century, biochemists now have the tools to study proteins under these more physiologically relevant conditions. We review a part of this burgeoning postreductionist landscape by focusing on high-resolution protein nuclear magnetic resonance (NMR) spectroscopy, the only method that provides atomic-level information over an entire protein under the crowded conditions found in cells.

Figure 1

Effects of large- and small- molecule crowders on the diffusion of CI2. The ratio of the diffusion coefficient in buffer to that in the co-solute is plotted for rotational and translational diffusion as a function of the relative macroscopic viscosity. The translational diffusion was measured by using the X-STE (92) pulse sequence, and rotational diffusion was measured from the ¹⁵N T₁/T₂ ratio (93). The experiments were performed on a 600 MHz spectrometer at 298 K in acetate buffer, pH 5.4.