Cell wall damage increases macromolecular crowding effects in the Escherichia coli cytoplasm

Abstract

The intracellular milieu is crowded with biomacromolecules. Macromolecular crowding changes the interactions, diffusion, and conformations of the biomacromolecules. Changes in intracellular crowding effects have been mostly ascribed to differences in biomacromolecule concentration. However, the spatial organization of these molecules should play a significant role in crowding effects. Here, we find that cell wall damage causes increased macromolecular crowding effects in the Escherichia coli cytoplasm. Using a genetically-encoded macromolecular crowding sensor, we see that crowding effects in E. coli spheroplasts and Penicillin G-treated cells well surpass crowding effects obtained using hyperosmotic stress. The crowding increase is not due to osmotic pressure, cell shape, crowder synthesis, or volume changes, and therefore not crowder concentration. Instead, a genetically-encoded nucleic acid stain and a small molecule DNA stain show nucleoid expansion and cytoplasmic mixing, which could cause these increased crowding effects. Our data demonstrate that cell stress from antibiotics or cell wall damage alters the biochemical organization in the cytoplasm and induces significant conformational changes in a probe protein.