Condition-dependent amorphous protein agglomerates control cytoplasmic rheology

Molecular crowding in the bacterial cytoplasm restricts diffusion of large molecules, impacting cellular processes. To monitor cytoplasmic diffusion and rheology, we used single-particle tracking in Escherichia coli, finding a 3-fold variation in the diffusion of a 40-nm particle across exponential growth conditions. Known determinants of rheology did not account for this variation. Instead, we found a strong anticorrelation between the diffusion coefficient and the abundance of amino acid metabolism proteins (clusters of orthologous groups [COG] category "E"), persisting upon genetic perturbations, and that lower diffusion is associated with increased elasticity. Photoactivated light microscopy revealed that some amino acid metabolism proteins form clusters. Electron microscopy showed that these proteins can form amorphous agglomerates at physiological concentrations in vitro due to their high hydropathy, which also confers low disorder and compactness. These findings show that diffusion is controlled by the formation of protein agglomerates and thus reveal how condition-induced proteome changes affect cytoplasmic rheology.