Macromolecular crowding tunes protein stability by manipulating solvent accessibility

Abstract

In all intracellular processes, protein structure and dynamics are subjected to the influence of macromolecular crowding (MC). Here, the impact of different types and sizes of MC agents on the model protein BsCspB are comprehensively investigated under thermal as well as chemical denaturation. We consistently reveal a distinct stabilization of BsCspB in dependence on the MC concentration but not on viscosity, polarity or size of MC agent used. This general stabilization is decoded by using NMR spectroscopy via monitoring chemical shift (CS) perturbations, the intramolecular hydrogen bonding network as well as local protection of amide protons against exchange with solvent protons. Whereas CSs and the hydrogen bonding network are not systematically affected in presence of MC, we detected a pronounced reduced exchange in loop regions of BsCspB. We conclude that this reduced accessibility of solvent protons acts as a key parameter for the increase in protein stability seen under MC.