Molecular crowding effects
Most of our general knowledge about biological chemistry or biochemistry stems from experiments on dilute samples of macromolecules, like proteins, or DNA, at concentrations of about a few grams per liter. However, these conditions are surprisingly different from those inside living cells. The cellular cytoplasm is crowded with many different macromolecules reaching concentrations with hundreds of grams per liter. Hence, a substantial fraction (20-30%) of the total volume of any cell is occupied by macromolecules. This so-called excluded volume increases the effective concentration of molecules and affects the physical properties of the solution (“excluded volume” effect). For example the molecular mobility and dynamics within constrained volumes are suppressed in the cytosol, characterized by a high concentration of large and small molecules. As a consequence molecular crowding decreases the diffusion rate, shifts the equilibrium of protein-protein association and of protein-substrate interactions and changes the conformational dynamics of proteins. We performed FCS studies on several proteins dissolved in highly concentrated solutions of Ficoll (of different molecular weights ranging from 5-400 kDa) which mimic molecular crowding.