T-jump and circular dichroism: folding dynamics in proteins and DNA

Abstract

We review recent experimental work combining circular dichroism spectroscopy and T-jump experiment to investigate elementary dynamics in the folding/unfolding of biomolecules. A nanosecond setup is presented for poly(Glutamic acid) studies whereas a millisecond one is developed for investigation of DNA G-quadruplex dynamics.

Folding Dynamics of DNA G-Quadruplexes Probed by Millisecond Temperature Jump Circular Dichroism

“…In standard experiments, powerful nanosecond lasers are generally used for the solvent excitation, which yielded a fast temperature rise that starts to decrease after ca. 1 ms due to the heat dissipation. , To extend the observation time window up to seconds, CW infrared (IR) excitation sources have been used in combination with single-molecule spectroscopy, microfluidics, and very recently 2D IR spectroscopy. ,, In a similar way, we implemented a 5 W CW laser diode emitting at 1.5 μm, for the sample excitation, in our T-jump setup . The IR beam was focused onto a 150 × 140 μm 2 spot in a 100 μm optical path sample cell to ensure the homogeneous heating and cooling of the sample by keeping a weak absorption of the solvent.…”

“…Unlike short DNA duplexes, the melting dynamics of short intramolecular G4s are expected to occur on longer time scales, , making pulsed laser T-jump initiation inappropriate for probing their dissociation. In this regard, recent developments have been achieved for the use of CW T-jump initiation to access both the denaturation and the renaturation dynamics of biomolecules beyond the millisecond time scale. − Herein, we present the first study of this type, devoted to G4-forming sequences. We compare the thermal denaturation and renaturation dynamics of four G4-forming sequences: three antiparallel topologies made of the human telomeric sequences Tel21 (d­[(5′-GGG­(TTAGGG) 3 -3′]) and Tel22 (d­[5′-AGGG­(TTAGGG) 3 -3′]) and the short thrombin binding aptamer sequence (d­[5′-GGTTGG­TGTGGTTGG-3′]) and the parallel topology made of the human CEB25 minisatellite sequence 26CEB (d­[5′-AAGGGTG­GGTGTAAGT­GTGGGTGGGT-3′]).…”

Multiscale conformational dynamics probed by time‐resolved circular dichroism from seconds to picoseconds

“…Switching on and off the IR beam is controlled by rapid shutters, allowing heating of the samples within a few tens of ms. 5 The experimental procedure, as observable in Figure 6A, is as follows: During the first second, there is no illumination, allowing the measurement of the steady-state CD, then IR light is switched on for 4 s, provoking a rapid heating of the sample and the subsequent denaturation of the G4. This effect translates into a decrease of the CD as observable in Figure 6A.…”

“…2,3 In this context, our group has been developing, for a few decades, complementary time-resolved CD (TRCD) techniques in order to investigate either ultrarapid (i.e., in the subpico-nanosecond time domain) 4 or slower (i.e., in microsecond up to the second time domain) processes in organic and inorganic compounds and in biomolecules. 5 Although local conformational changes that are restricted to a few atoms generally occur on the ultrarapid timescale, more global phenomena, such as the changes of protein or DNA secondary structure, occur on much longer timescales.…”