Closed loop folding units from structural alignments: experimental foldons revisited.

Nonoverlapping closed loops of around 25-35 amino acids formed via nonlocal interactions at the loop ends have been proposed as an important unit of protein structure. This hypothesis is significant as such short loops can fold quickly and so would not be bound by the Leventhal paradox, giving insight into the possible nature of the funnel in protein folding. Previously, these closed loops have been identified either by sequence analysis (conservation and autocorrelation) or studies of the geometry of individual proteins. Given the potential significance of the closed loop hypothesis, we have explored a new strategy for determining closed loops from the insertions identified by the structural alignment of proteins sharing the same overall fold. We determined the locations of the closed loops in 37 pairs of proteins and obtained excellent agreement with previously published closed loops. The relevance of NMR structures to closed loop determination is briefly discussed. For cytochrome c, cytochrome b(562) and triosephophate isomerase, independent folding units have been determined on the basis of hydrogen exchange experiments and misincorporation proton-alkyl exchange experiments. The correspondence between these experimentally derived foldons and the theoretically derived closed loops indicates that the closed loop hypothesis may provide a useful framework for analyzing such experimental data.