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Spectroscopie infrarouge impulsionnelle appliquée au transfert de ligands dans les hémoprotéines

Abstract : Ligand transfer (of diatomics such as O2, CO, NO, CN...)in heme proteins is an integral part of many biological functions such as storage and ligand transport, enzymatic catalysis and ligand detection. A thorough knowledge of these mechanisms underlying the ligand transfer from the heme binding site to the protein exterior is essential to understand how these heme proteins accomplish their function : discriminate between ligands, shuttle them from the solvent to the heme pocket and vice versa, and react to external signals.

The ligand transfer from the heme binding site to the exterior of the protein involves different stages with timescales spanning several orders of magnitude. Our study deals with the very first events of the ligand transfer which occur on the femtosecond timescale. In the case of myoglobin case, this event is the transfer of the ligand from the heme to a nearby site called docking-site, which is the first site the ligand reaches on its way to the solvent.

We monitor this transfer using infrared (5 µm) femtosecond pulses which directly probe the CO ligand vibrational changes during the transfer. The vibrational dephasing time of the CO ligand (1 picosecond) is longer than the dynamics of the transfer. Therefore, coherent effects arise and forbid a direct interpretation of spectrally resolved transmission experiments at short pump-probe time delays. We avoid these effects by using two complementary methods. In a first experimental configuration, the transmission of the sample is spectrally integrated, we are thus insensitive to coherent effects. In such a scheme, a very sensitive detection is required in order to access the weak differential signal. In a second experimental configuration, we monitor the ligand vibration during the transfer by recording the emitted field of the CO. The homodyne detection of this field requires phased-locked mid-IR pulses.

We developed a non-stationary response function which describes the sample response and allows the simulation of these two kinds of experiments. We also introduce a novel time-frequency representation of the non-stationary response function.

We performed experiments on myoglobin, a reference system for the ligand transfer study. In this protein, spectrally integrated differential transmission experiments showed a progressive
decay of the oscillator strength of the ligand during the transfer. Simulations based on a phenomenological model support this interpretation. Surprisingly, the oscillator strength
change does not follow the quasi-instantaneous vibrational frequency change. To our knowledge, this is the first report on a progressive decay of the vibrational oscillator strength following the breaking of a chemical bond. This oscillator strength correlated to heme-ligand distance is a potential probe of the ligand transfer.
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Contributor : Thomas Polack <>
Submitted on : Monday, March 8, 2004 - 7:09:05 PM
Last modification on : Monday, October 19, 2020 - 10:59:59 AM
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  • HAL Id : tel-00005255, version 1



Thomas Polack. Spectroscopie infrarouge impulsionnelle appliquée au transfert de ligands dans les hémoprotéines. Physique Atomique [physics.atom-ph]. Ecole Polytechnique X, 2003. Français. ⟨tel-00005255⟩



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