Waltzing alpha-helices.

P roteins are 30% -helices, which, together with -sheets and loops, self-assemble into specific topological arrangements that make biologically active 3dimensional structures. The -helix has another important feature: it is capable of folding autonomously (1). Despite the apparently simpler structure, -helix formation is governed by the same physical principles as protein folding, and recruits a similar array of interactions for its stabilization, including hydrogen bonds, electrostatics, dipole–dipole, and hydrophobic interactions (2). Furthermore, isolated -helices display very complex conformational behavior. All of these properties have made the -helix an excellent test lab for protein-folding research. From such efforts we now understand the factors that determine -helix stability (3) and the timescales and mechanism of -helix formation (4). New nanosecond laser-induced temperature-jump techniques can detect the kinetics of individual residues within the -helix (5), producing exciting data with which to refine our understanding of helix formation. However, what has been missing is a technique to detect the complex motions that should take place in the nanosecond timescale in isolated -helices. In an article appearing in a recent issue of PNAS, Fierz et al. (6) describe the application of the contact formation ultrafast kinetic technique to monitor nanosecond conformational f luctuations in -helices at equilibrium conditions. The method promises to directly report on previously unobserved and important conformational processes of already formed helical segments, such as motion resulting from helix melting at one end and growth at the other. In the theoretical description of helix formation, already developed in the late 1950s (7), short helical segments are thought to be highly unstable and thus relatively slow to form, whereas growth into longer helices is both more favorable and much faster. Pioneering experiments on very long nonnatural homopolymers quickly followed (8), but it took another 2 decades to develop the tools to study short protein-like -helices. The discovery of the first protein segment capable of forming helix structure on its own (1) and the development of simple design principles to produce short helical peptides resulted in extensive experimental characterization of -helix stability (3). From these experiments, we learned that nucleation is only slightly unfavorable. In other words, helix formation is weakly cooperative, resulting in complex distributions of helical segments of different size and position along the peptide sequence. Such empirical data from hundreds of designed and natural peptides were introduced into nucleation–elongation models producing a theory with real predictive power, demonstrating that the general principles behind -helix stability were well understood (9). Helix kinetics studies followed on. The timescales of -helix formation were among the first phenomena measured with ultrafast folding techniques, most notably the laser-induced temperature jump (10, 11). The overall timescale for the relaxation between coil and helices was discovered to be hundreds of nanoseconds (10, 12). Analysis with a kinetic nucleation–elongation model demonstrated consistency between Tjump kinetic experiments and the nucleation barrier required for the equilibrium data, and estimated an elementary rate of helix propagation of 1–4 ns per residue (12). Kinetic theory made another intriguing prediction. In addition to the overall relaxation corresponding to motion over the nucleation barrier, there should be a fast process corresponding to interconversion among preformed helical segments. The fast process would arise from packets of propagation-shortening events and thus should take longer than the 1to 4-ns single-residue rotations, but shorter than the overall equilibration with the coil. The problem is that this fast process was not observed in infrared (10) nor in fluorescence (12) T-jump experiments. The underlying kinetic complexity of helix formation became apparent when laser T-jump experiments were combined with infrared detection of single residues by using specific isotopic labeling (5, 13). But again, detailed theoretical analysis, this time including the complete description of -helix stability and ac-