For over fifty years, the unfolded state of proteins had been thought to be
featureless and random. Experiments by Tanford and Flory confirmed that unfolded
proteins possessed the same dimensions as those predicted of a random flight chain in
good solvent. In the late eighties and early nineties, however, researchers began to notice
structural trends in unfolded proteins. Some experiments showed that the unfolded state
was very similar to the native state, while others indicated a conformational preference
for the polyproline II helix in unfolded proteins. As a result, a paradox developed. How
can unfolded proteins be both random and nonrandom at the same time?
Current experiments and most theoretical simulations cannot characterize the
unfolded state in high detail, so we have used the simplified hard sphere model of
Richards to address this question. By modeling proteins as hard spheres, we can not only
determine what interactions are important in the unfolded state of proteins, but we can
address the paradox directly by investigating whether nonrandom behavior is in conflict
with random coil statistics.
Our simulations identify hundreds of disfavored conformations in short peptides,
each of which proves that unfolded proteins are not at all random. Some interactions are
important for the folded state of proteins as well. For example, we find that an α-helix
cannot be followed directly by a β-strand because of steric considerations. The
interactions outlined here limit the conformational possibilities of an unfolded protein far
beyond what would be expected for a random coil. For a 100-residue protein, we find
that approximately 9 orders of magnitude of conformational freedom are lost because of
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local chain organization alone. Furthermore, we show that the existence of this
organization is compatible with random coil statistics.
Although our simulations cannot settle the controversy surrounding the unfolded
state, we can conclude that new methods of characterizing the unfolded state are needed.
Since unfolded proteins are not random coils, the methods developed for describing
random coils cannot adequately describe the complexities of this diverse structural
ensemble