Oxygen depletion in biomolecular condensates is dominated by macromolecular density.

Biomolecular condensates are dynamic cellular compartments formed by the self-assembly of proteins and nucleic acids. Many metabolites partition into condensates based on their interactions with the macromolecular constituents; yet, whether gases behave similarly remains unknown. Here, we show that oxygen partitions into protein-based condensates formed by intrinsically disordered repeat proteins with systematically varied sequences. Using microelectrodes, phosphorescence lifetime imaging microscopy, and molecular dynamics simulations, we find that oxygen is partially excluded from the condensate, and its partitioning does not correlate with the condensate hydrophobicity. Instead, oxygen concentration is inversely related to condensate protein density. These results suggest that the prevailing theory of small-molecule partitioning into condensates has to be augmented to consider the concentration of macromolecules as a dominant factor in the absence of interactions between the metabolite and the condensate. Our results suggest that biomolecular condensates can generate a nanoscale oxygen gradient, potentially modulating the local availability of oxygen for biochemical reactions within the cell.