Effects of macromolecular crowding on protein folding and aggregation

We have studied the effects of polysaccharide and protein crowding agents o n the refolding of oxidized and reduced hen lysozyme in order to test the p rediction that association constants of interacting macromolecules in livin g cells are greatly increased by macromolecular crowding relative to their values in dilute solutions. We demonstrate that whereas refolding of oxidiz ed lysozyme is hardly affected by crowding, correct refolding of the reduce d protein is essentially abolished due to aggregation at high concentration s of crowding agents. The results show that the protein folding catalyst pr otein disulfide isomerase is particularly effective in preventing lysozyme aggregation under crowded conditions, suggesting that crowding enhances its chaperone activity. Our findings suggest that the effects of macromolecula r crowding could have major implications for our understanding of how prote in folding occurs inside cells.