Fast folding dynamics of α-helical peptides – Effect of solvent additives and pH

Measurement of circular dichroism dynamics in a nanosecond temperature-jump experiment

“…For verification of our results, we performed T-jump experiments detected at 1632 cm −1 using a setup which has been described in detail before. 8 Briefly, the solvent (D 2 O) was heated using ns-laser pulses at 1970 nm, generated by a Nd:YAG/dye laser system with IR difference frequency generation. Absorbance changes at 1632 cm −1 were probed using the continuous IR beam from a tunable lead-salt diode laser and a fast HdCdTe photodiode with 50 MHz bandwidth, yielding an overall signal rise time of 14 ns.…”

“…5, the temperature remains constant over more than 800 µs, much longer than the 200 µs timescale observed for cells of comparable pathlength with CaF 2 windows. 8 This can be an advantage for measuring longer processes. Furthermore, we did not observe any cavitation effects, a convenient feature for experiments.…”

“…Alternatively, one can directly heat the water solvent using IR laser pulses. The latter method yields less cavitation problems, 8 but demands a nanosecond IR source. Most T-jump experiments based on an IR probe necessitate the use of deuterated water and require IR pulses at 1.9 µm.…”

“…Furthermore, we did not observe any cavitation effects, a convenient feature for experiments. We think that this is a consequence of the direct heating (using IR laser pulses), which allows measurements in the absence of dye molecules which tend to aggregate and act as nucleation sites for cavitation, 8 and of the smooth pump beam profile employed here, which prevents the occurrence of hot spots leading to the onset of cavitation. Under these conditions, cavitation can be avoided, even at elevated base temperatures.…”

“…4 Second, it is well known that the IR-Amide I' band is sensitive to the secondary structure of proteins. 5 This feature has been extensively studied in conjunction with T-jumps to investigate unfolding dynamics of model polypeptides [6][7][8] or small proteins. 9,10 However, these techniques suffer from a lack of quantitative information.…”