Formation of Protein-Complexes in crowded environments: From in vitro to in vivo
Prof. Gideon Schreiber, Dept. Biological Chemistry, Weizmann Institute of Science
Rates of protein interactions are one to five orders of magnitude slower than the theoretically calculated collision rate of spheres of the same size. The rates can be increased by favorable electrostatic forces between the two proteins. While a large number of studies have been done in the test tube, much less is known about the effect of the crowded complex environment in the cell on interaction rates and affinities. Here, I will discuss our experimental results on association in living cells and in other crowded environments, and put them into perspective of association studies performed in the test tube. In recent years, we and others have established that the association reaction proceeds through transient complexes, which may be specific or diffusive in nature. To bring binding studies closer to the in vivo environment, we investigated the role of crowding and the cellular environment on binding. For crowding we added various polymers to the solution, including Dextran and PEGs of different molecular weights, used cell extracts or performed real time measurements within living cells. While crowding enhances oligomerization and polymerization of macromolecules, it has only a small effect on the binding rates and affinities of transient protein-protein interactions. We suggest that the limited effect of crowders, which is much bellow the expected from the increased viscosity of the solutions, is a result of the occluded volume effect in high crowder concentrations. Direct measurements of the stability of the encounter complex shows that crowders have only a minor effect on K1 or k2, and do not change the shape or size of the encounter complex. The results will be discussed in light of the occluded volume effect of crowders. The results suggest that typical crowding agents have only a small effect on specific protein-protein dimerization reactions while promoting aggregation.