  The folding dynamics of small single-domain proteins is a current focus of
simulations and experiments. Many of these proteins are 'two-state folders',
i.e. proteins that fold rather directly from the denatured state to the native
state, without populating metastable intermediate states. A central question is
how to characterize the instable, partially folded conformations of two-state
proteins, in particular the rate-limiting transition-state conformations
between the denatured and the native state. These partially folded
conformations are short-lived and cannot be observed directly in experiments.
However, experimental data from detailed mutational analyses of the folding
dynamics provide indirect access to transition states. The interpretation of
these data, in particular the reconstruction of transition-state conformations,
requires simulation and modeling. The traditional interpretation of the
mutational data aims to reconstruct the degree of structure formation of
individual residues in the transition state, while a novel interpretation aims
at degrees of structure formation of cooperative substructures such as
alpha-helices and beta-hairpins. By splitting up mutation-induced free energy
changes into secondary and tertiary structural components, the novel
interpretation resolves some of the inconsistencies of the traditional
interpretation.
