Protein and DNA folding dynamics probed by time-resolved circular dichroism

Protein and DNA folding dynamics probed by time-resolved circular dichroism
© Ecole polytechnique


Pascale Changenet
Francois Hache
Auriane Perrin (PhD)

Internship proposal

Research topics:

Besides their 3D structure, the dynamical properties of biomolecules play a key role in their functions. During their folding, they may explore a rich set of conformations in the fold space that may be difficult to capture experimentally. In this regard, chiroptical methods are valuable tools for investigating their structural changes over large time windows. Among them, circular dichroism (CD) has become very popular with the advantage that the CD spectrum of biomolecules can be theoretically or phenomenologically related to their conformations. Time-resolved experiments remain however technically challenging, due to their extremely low signals.

Our main activity focusses on the development and the use of new “user-friendly” chiroptical tools for measuring the conformational dynamics of proteins and DNA, over a time scale spanning femtoseconds to seconds. Part our activity is also devoted to the study of the conformational dynamics of small chiral organic compounds in their excited state. 


Ongoing projects:

  • Experimental developments


In these last years, we have built two complementary monochromatic TRCD set-ups allowing either to monitor the conformational changes of biomolecules following femtosecond optical excitation with a sub-picosecond time resolution or to follow their thermal denaturation (T-jump) on the nanosecond up to the millisecond timescale. We have recently developed a novel extension of our T-jump/TRCD set-up capable to measure the thermal denaturation and the consecutive renaturation of DNA over a time window spanning a few ten milliseconds up to seconds.

  • Folding mechanism of DNA G-quadruplexes


DNA can adopt a large variety of conformations. Among them, G-quadruplexes (G4) are non-canonical DNA structures that result from the hydrophobic stacking of a number of guanine quartets in the presence of metallic cations such as K+ or Na+. After intense speculations about their biological pertinence, there is now compelling experimental evidence of their participation in important cellular regulation functions correlated to their folding mechanism. While there have been numerous studies of the 3D structure of G4, mechanisms of formation are only partially understood. This project aims at investigating the folding dynamics of short single-stranded G4 structures, on a time window spanning from picoseconds to seconds with a combination of TRCD spectroscopies. We have measured the denaturation and the consecutive renaturation kinetics of several antiparallel G4. These first results suggest the existence of multiple folding routes along extremely rugged energy landscapes that strongly depend on the applied perturbation and the concentration of metallic cations. Comparative studies with other G4 topologies are currently underway in collaboration with the synchrotron DISCO-SOLEIL.        

  • Chirality control in small organic compounds

Control of chirality both at molecular and macroscopic levels is an important issue of the contemporary chemistry. In this regard, axially chiral molecules, such as 1,1′-bi-2-naphthol (binol), have been attracting a great deal of interest for several decades. Binols have been widely used as chiral templates for a broad range of applications in molecular recognition and asymmetric catalysis. More recently, their use has been extended to the design of functional chiral supramolecular assemblies and innovative organic materials for applications in linear and non-linear optics. Chirality of binols arises from the hindered rotation of their two naphthyls subunits providing them strong chiroptical activity. While significant efforts have been done to understand the origin of the chiroptical properties of hindered binaphthyls in their ground state, their fate in their excited state is poorly understood.



Owing to the recent improvements of our sub-picosecond TRCD set-up, we have been able to perform the first exhaustive study of three bridged binols derivative in various solvents. These measurements allowed us to disentangle the respective effects of the solvent and the substituents on the chiroptical properties of hindered binaphthyls in their excited-state. In particular, we have shown that, the presence of the bridges does not prevent the conformational changes of binol in their excited state leading to a fast and significant increase of their CD response.



Frank Wien, Matthieu Réfrégiers, DISCO-SOLEIL, Saint Aubin.
Bastien Nay, LSO, Ecole Polytechnique, Palaiseau.
Brahim Heddi, Laboratoire de Biologie et Pharmacologie Appliquée, ENS Paris-Saclay.
Dimitra Markovitsi, Thomas Gustavsson, LIDYL, CEA Saclay.
Roberto Improta, Istituto Biostrutture e Bioimmagini, Consiglio Nazionale delle Ricerche, Napoli, Italy.
Fabrizio Santoro, Consiglio Nazionale delle Ricerche, Istituto di Chimica dei Composti Organometallici, Pisa, Italy.
Stephan Guy, Institut Lumière Matière, Lyon.

Selected publications:

Artifact-free balanced detection for the measurement of circular dichroism with a sub-picosecond time resolution
P. Changenet and F. Hache, Opt. Expr.

Multiscale conformational dynamics probed by Time-resolved circular dichroism from milliseconds to picoseconds
F. Hache and P. Changenet, Chirality ,

Folding dynamics  of DNA G-quadruplexes probed by millisecond T-jump circular dichroism
K. Laouer, M. Schmid, F. Wien, P. Changenet, F. Hache,  J. Phys. Chem. B, 125, 8088 (2021)

Introduction to Femtochemistry: Excited-state Proton Transfer from Pyranine to Water Studied by Femtosecond Transient Absorption
P. Changenet, T. Gustavsson, I. Lampre,  J. Chem. Edu., 97, 4482 (2020)

Unveiling excited-state chirality of Binaphthols by sub-picosecond circular dichroism and quantum-chemical calculations
P. Changenet, F. Hache, SPIE  11277, Article Number: 112770W (2020)

Transient absorption in inorganic systems
M. Ha-Thi, G. Burdzinski, T. Pino, P. Changenet, Springer Handbook of Inorganic Photochemistry, Accepted

A comprehensive study of guanine excited state relaxation and photoreactivity in G-quadruplexes
L. Martinez-Fernandez, P. Changenet, A. Banyasz, T. Gustavsson, D. Markovitsi, R. Improta,  J. Phys. Chem. Lett. 10, 6873 (2019)

T-jump and circular dichroism: Folding dynamics in proteins and DNA
P. Changenet, F. Hache,  Mol. Cryst. Liq. Cryst. 693, 49 (2019)

Unveiling excited-state chirality of binaphthols by femtosecond circular dichroism and quantum-chemical calculations
M. Schmid, L. Martinez-Fernandez, D. Markovitsi, F. Santoro, F. Hache, R. Improta, P. Changenet, J. Phys. Chem. Lett. 10, 4089 (2019)

Ultrafast Electron Transfer in Complexes of Doxorubicin with Human Telomeric G-Quadruplexes and GC Duplexes Probed by Femtosecond Fluorescence Spectroscopy
P. Changenet-Barret, T. Gustavsson, D. Markovitsi, I. Manet, ChemPhysChem 17, 1264 (2016)

Ultrafast conformational changes in biomolecules studied by time-resolved circular dichroism
P. Changenet-Barret, F. Hache, SPIE  9652, Article Number: 96520Z (2015)

Comparative study of the folding/unfolding dynamics of poly(glutamic acid) in light and heavy water
L. Mendonça, A. Steinbacher, R. Bouganne and F. Hache, J. Phys. Chem. B 118, 5350 (2014)

Ultrafast carbonyl motion of the Photoactive Yellow Protein probed by femtosecond circular dichroism 
L. Mendonça, F. Hache, P. Changenet-Barret, P. Plaza, H. Chosrowjan, S. Taniguchi and Y. Imamoto, J. Am. Chem. Soc. 135, 14637 (2013)