FAST EVENTS IN PROTEIN FOLDING: The Time Evolution of Primary Processes

Abstract

Most experimental studies on the dynamics of protein folding have been confined to timescales of 1 ms and longer. Yet it is obvious that many phenomena that are obligatory elements of the folding process occur on much faster timescales. For example, it is also now clear that the formation of secondary and tertiary structures can occur on nanosecond and microsecond times, respectively. Although fast events are essential to, and sometimes dominate, the overall folding process, with a few exceptions their experimental study has become possible only recently with the development of appropriate techniques. This review discusses new approaches that are capable of initiating and monitoring the fast events in protein folding with temporal resolution down to picoseconds. The first important results from those techniques, which have been obtained for the folding of some globular proteins and polypeptide models, are also discussed.

Conformational changes during the nanosecond-to-millisecond unfolding of ubiquitin

“…The T-jump before transient DVE probing is created by nanosecond excitation of the OD stretch overtone of the D 2 O buffer solution at ϭ 2 m. Picosecond vibrational relaxation raises the temperature of the solution on the time scale of the excitation pulse (15). The 6-mJ, 8-ns T-jump pulse is obtained from a beta barium borate (BBO)-based optical parametric oscillator pumped by the output of a 20-Hz, frequency-doubled, Q-switched Nd:yttrium͞aluminum-garnet laser and focused to a 500-m diameter at the sample.…”

“…Optical and IR spectroscopies such as fluorescence-based methods (10,11), time-resolved circular dichroism (12), Raman (13), and IR absorption (14,15) are desirable in fast folding experiments because they have the intrinsic time resolution to follow the fastest processes. Nevertheless, relating these experiments to nuclear coordinates that characterize a protein structure or conformation is not trivial, particularly in the presence of disorder.…”

“…IR spectroscopy on amide I transitions (primarily CO stretching; 1,600-1,700 cm Ϫ1 ) has been widely used for folding studies of proteins (7,(14)(15)(16)(17) and peptides (9,(18)(19)(20), primarily because amide I spectra have peak positions that depend on secondary structure. The frequency-structure correlation is a result of through-space electrostatic couplings that depend on the distance and orientation between the many amide I vibrations of each of the protein's peptide units (21,22).…”

Vapor Treatment of Electrospray Droplets: Evidence for the Folding of Initially Denatured Proteins on the Sub-Millisecond Time-Scale

“…Knowledge about folding transitions can aid in the understanding of biological activity, as biological activity is often regulated by the higher order structure [1]. However, the lifetimes of many partially folded species are usually on the order of hundreds of microseconds to fractions of a second, thus making it difficult to study these intermediates using X-ray or NMR methods [2]. With the development of electrospray ionization (ESI), many mass spectrometry-based methodologies have been established to elucidate the higher order structure of biomolecules.…”

Stepwise helix formation and chain compaction during protein folding

“…Temperature jump and other rapid perturbation methods have shown that isolated helices and ␤-hairpins form and decay over a time window ranging from Ϸ100 ns to several microseconds (4,5). Recent advances in rapid mixing techniques combined with structurally informative spectroscopic probes made it possible to resolve conformational events on the submillisecond time scale preceding the rate-limiting step in the folding of globular proteins (6)(7)(8)(9).…”