  Here, we show by solution nuclear magnetic resonance measurements that the
urea-unfolded protein apoazurin becomes elongated when the synthetic crowding
agent dextran 20 is present, in contrast to the prediction from the
macromolecular crowding effect based on the argument of volume exclusion. To
explore the complex interactions beyond volume exclusion, we employed
coarse-grained molecular dynamics simulations to explore the conformational
ensemble of apoazurin in a box of monodisperse crowders under strong chemically
denaturing conditions. The elongated conformation of unfolded apoazurin appears
to result from the interplay of the effective attraction between the protein
and crowders and the shape of the crowders. With a volume-conserving crowder
model, we show that the crowder shape provides an anisotropic direction of the
depletion force, in which a bundle of surrounding rod-like crowders stabilize
an elongated conformation of unfolded apoazurin in the presence of effective
attraction between the protein and crowders.
