Probing Electrostatic\nand Hydrophobic Associative\nInteractions in Cells

Weak nonspecific\ninteractions between biomacromolecules determine\nthe cytoplasmic organization. Despite their importance, it is challenging\nto determine these interactions in the intracellular dense and heterogeneous\nmixture of biomacromolecules. Here, we develop a method to indicate\nelectrostatic and hydrophobic associative interactions and map these\ninteractions. The method relies on a genetically encoded probe containing\na sensing peptide and a circularly permuted green fluorescent protein\nthat provides a ratiometric readout. Inside bacterial and mammalian\ncells, we see that the cytoplasmic components interact strongly with\ncationic and hydrophobic probes but not with neutral hydrophilic probes,\nwhich remain inert. The Escherichia coli cytoplasm interacts strongly with highly negatively charged hydrophilic\nprobes, but the HEK293T cytoplasm does not. These associative interactions\nare modulated by ATP depletion. Hence, the nonspecific associative\ninteraction profile in cells is condition- and species-dependent.