Towards a molecular movie: Transient 2D-IR spectroscopy Peter Hamm Physikalisch Chemisches Institut, Universität Zürich

B.L. de Groot and H. Grubmüller, Science 294, (2001) Biosystems are Dynamical Aquaporin-1

Structure Sensitive Methods Scattering Methods (x-ray, electrons, neutrons) 2D-NMR + highest structure resolution power - works best in crystals + works best in solution + high structure resolution power + intrinsic time resolution unlimited - intrinsic time resolution very poor (1-100 ms) Time Resolved Scattering Methods 2D-IR + intrinsic time resolution fast (1 ps) - structure resolution power low but non-zero

Site BasisExciton Basis Coupled Vibrational States Linear Coupling:

Frequency C N H C O C N H C O C N H C O H H H C N H C O R1R1 C N H C O R3R3 C N H C O R2R2 Weak coupling, heterogeneous Strong coupling, homogeneous R2R2 R3R3 R1R1 Linear Spectroscopy

t p =100 fs  =200 cm -1 E=1 µJ Spectrometer + 2  32 Element MCT Detector Array Fabry Perot Filter Delay Sample 2D-IR Spectroscopy: Setup

2D-IR Spectroscopy |0  |1  |2  |0‘  |1‘  |2‘ 

2D-IR Spectroscopy |10  |20  |11‘  |00‘  |01‘  |02‘ 

2D-IR Spectroscopy: Equilibrium Woutersen et al. J. Phys. Chem. B, 104, (2000) Woutersen et al., PNAS 98, (2001) Woutersen et al. J. Chem. Phys. 117, (2002) Probe Frequency [cm -1 ]  Conformation and distribution of conformations  Ultrafast equilibrium fluctuations

Our Dream: Molecular Movie

Photocleavable Peptide C. Kolano, K. Gomann, W. Sander, Eur. J. Org. Chem. 2004, 20, h ?

Photocleavable Peptide: IR spectrum Boc-Cys-Pro-Aib-Cys-OMe

Equilibrium 2D Spectrum 1 ps t 2D-IR

Equilibrium 2D Spectrum 1 ps t 2D-IR

Equilibrium 2D Spectrum 1 ps t 2D-IR

IR Probe Spectrometer + 2  32 Element MCT Detector Array Delay Sample Flow-Cell Transient 2D Spectroscopy: Setup UV Switch Delay Fabry Perot Filter IR Pump

Transient 2D-IR Spectra ps1 ps t IR pump IR probe UV pump 3 ps25 ps100 ps

MD Package: Gromacs; Gromos Force Field 43a1 Starting geometry: Based on low-resolution x-ray structure Solvent box: Acetonitrile (1048 molecules) Nonequilibrium MD Simulations Equilibrium Simulation (NVT): 120 ns 1 ns Bondbreaking: Remove S-S Bond Non-Equilibrium Simulation (NVE): 600 ps

MD Simulations: Example Trajectories 3.0 Å

MD Simulations: Average 3.0 Å 6.1 Å ++ --

Transient 2D-IR Spectra Equilibrium 2DTransient 2D 100 ps C. Kolano, Nature 2006 in press

Transient 2D-IR Spectra Equilibrium 2DTransient 2D 100 ps

Transient 2D-IR Spectra Equilibrium 2DTransient 2D 100 ps

And Finally: The Movie

Conclusion  Transient 2D-IR spectroscopy combines picosecond time resolution with microscopic structural information  Transient 2D-IR reveals new insight into fast structural dynamics of peptides, that cannot be obtained otherwise.  An ultrafast molecular movie of the 3D- structure conceivable (in connection with isotope labeling)

Outlook: Isotope Labelling h 13 C=O 13 C= 18 O  -Helix Folding

University ZürichUniversity Munich - Dr. Christoph Kolano - Prof. Wolfgang Zinth - Dr. Jens Bredenbeck - Dr. Jan Helbing University Bochum - Mariusz Kozinski - Dr. Christoph Kolano - Valentina Cervetto- Prof. Wolfram Sander - Harald Bregy - Ellen Backus Max Planck Institut, Martinsried - Virgiliu Botan - Dr. Christian Renner - Prof. Luis Moroder Max Born Institut, Berlin - Dr. Sander Woutersen University of Toronto - Prof. Thomas Elsaesser - Prof. Andrew Woolley University Frankfurt Financial Support - Prof. Josef Wachtveitl- Universität Zürich - Prof. Gerhards Stock - Swiss Science Foundation - Deutsche Forschungsgemeinschaft Acknowledgement