To come... For the moment please refer to these overview articles:
 J. Bredenbeck, Nachr. Chem. 54, 104-108 (2006) (in German)
labels provide new observables for protein structure and dynamcis
Time-resolved 1D and 2D-IR spectroscopy are powerful tools to study structure and dynamics of small to medium size molecules. When applied to proteins, these techniques face similar difficulties as known from classical FTIR spectroscopy: The protein spectrum contains many overlapping absorptions of the protein backbone (amide I and amide II modes) and the side chains. IR spectroscopy of proteins therefore frequently suffers from a lack of structural resolution.
In our group we combine time-resolved IR-spectroscopy with the aplication of artificial amino acids featuring functional groups that are not commonly present in proteins. Their infrared absorptions appear in a spectral window (ca. 1800 cm-1 to 3000 cm-1) that is free of native protein absorptions. These infrared labels therefore can provide site-selective information on the single amino acid level. Of special interest are nitriles and azides that can be incorporated cotranslationally by genetic code expansion or as a surrogate of a natural amino acid.
Solid lines are measured spectra of H2O (brown) abd D2O (blue).
Schematic IR spectrum of a protein (blue) and infrared labels (red).
Methods, Keywords: biochemistry, molecular biology, artificial amino acid, non-canonical amino acids, artificial genetic code expansion, codon reassignment, FTIR spectroscopy, time-resolved (2D-)IR spectroscopy, CD spectroscopy, ITC (isothermal titration calorimetry), DFT computations, molecular docking, protein structure visualization
of novel pulse
The dynamics of proteins is characterized by a continuum of time scales extending from the femtosecond regime of local dynamics involving just a few chemical bonds to the millisecond and longer time scale of correlated global structural changes. How do processes on such different time and length scales couple to each other, e. g. how is an ultrafast local change at the active site or binding site of a protein amplified into a mesoscopic change of structure that executes a biological function?
To gain access to such a wide range of time scales, we rely on electronically synchronized femtosecond lasers. Traditional femtosecond spectroscopy uses a trigger pulse and a detection pulse that are derived from the same parent pulse of the laser system, simply using partially reflective mirrors. Trigger pulse and measurement pulse are delayed with respect to each other using a delay line with motorized mirrors. Delays that can be achieved with an optical delay line typically are in the range of femtoseconds to 1 ns.
Fig.: Home built femtosecond laser synchronization.
Each of the two synchronized laser systems consists of an oscillator and an amplifier. One system serves as the master and the other as slave. One system provides the trigger pulse and the other the detection pulse. In this way arbitrary delays can be generated. To achieve synchronisation, the cavity length of the slave oscillator is continuously adjusted to match the master oscillator cavity. A phase-looked loop is used to force the difference frequency of both lasersystems to zero. The jitter can be as low as a few 100 fs. An electronic phase shifter generates delay between the oscillator pulses. Longer delays are created by picking different oscillator pulses for amplification in each of the amplifiers. This allows for pump-probe delays up to the repetition time of the laser systems of 1 ms.
and reaction mechanisms
Determining the structure of reactive intermediates is the key for understanding chemical reaction mechanisms and for developing and optimizing reactions. To resolve intermediate structures, a method combining structure sensitivity and high time resolution is required. Ultrafast polarization-dependent 2D-IR (P2D-IR) spectroscopy is an excellent complement to commonly used methods such as one-dimensional IR and multidimensional NMR spectroscopy for investigating reaction intermediates . P2D-IR allows structure determination by measuring the angles between vibrational transition dipole moments and by determining anharmonic couplings. Its high time resolution makes P2D-IR spectroscopy an attractive method for structure determination in the presence of fast exchange and for short-lived intermediates. The ubiquity of vibrations in molecules ensures broad applicability of the method, particularly in cases in which NMR spectroscopy is challenging due to a low density of active nuclei [1-3].
P2D-IR spectroscopy resolves reactive intermediates of Lewis acid catalyzed Diels-Alder reactions.
A minor conformer, source of unwanted side reactions, is revealed .
A. T. Messmer, K. M. Lippert, S. Steinwand, E.-B. W. Lerch, K. Hof, D.
Ley, D. Gerbig, H. Hausmann, P. R. Schreiner, J. Bredenbeck, Chem. Eur.
J. 18, 14989-14995 (2012)
Starting and stopping chemical reactions by light offers ultimate spatiotemporal control. Photocontrolled catalysts allow to initiate chemical reactions, which are no photochemical reactions themselves, by light. The figure shows the prototype of a reversibly switchable azobenzene-based catalyst. Isomerization of the azobenzene moiety activates the catalyst by exposing the catalytically active site. We study the dynamics of this and related catalysts as well as their interaction with the surrounding solvent and substrate molecules.
Reversibly photoswitchable catalyst .
In the inactive form, the active site is shielded by bulky rests R' on the azobenzene.
 M. V. Peters, R. S. Stoll, A. Kühn, S. Hecht, Angew. Chem. Int. Ed. 47, 5968-5972 (2008).
|Ultrafast 2D-IR spectroelectrochemistry
Electron transfer reactions are fundamental processes in chemistry and biology. They are key events in cellular respiration, photosynthesis and catalysis. We introduced the technique of 2D-IR spectroelectrochemistry to be able to carry out 2D-IR experiments under in situ redox control . The cell design allows for rapid redox cycling which is of particular importance for collecting redox induced 2D-IR difference spectra to measure subtle changes in structure and dynamics.
2D-IR spectroelectrochemistry is carried out in reflection mode
on a gold mirror which is used at the same time as an electrode.[1,2]
 Y. El Khoury, L. J. G. W. van Wilderen, J. Bredenbeck, Ultrafast 2D-IR spectroelectrochemistry of flavin mononucleotide, J. Chem. Phys. 142, 212416 (2015).
of natural and designed
The dynamics of proteins is characterized by a continuum of time scales extending from the femtosecond regime of local dynamics involving just a few chemical bonds to the microsecond and longer time scale of correlated global structural changes. How do processes on such different time and length scales couple to each other? How is an ultrafast local change at the active site or binding site of a protein amplified into a mesoscopic change of structure that executes a biological function? Related phenomena play a role in light sensitive proteins, where local ultrafast conformational changes of a chromophor drive processes on much larger time and length scales. Understanding the interaction of various degrees of freedom is of particular importance when it comes to the design of proteins. To study these macromolecular machines at work over many orders of magnitude in time with high structure resolution, we apply our tools of laser synchronization, multidimensional spectroscopy and incorporation of infrared labels by artificial amino acids.
Green fluorescent protein optimized for charge separation using
a genetically encoded metal-chelating amino acid to create an artificial copper redox center .
 X. Liu, J. Li, J. Dong, C. Hu, W. Gong, J. Wang, Angew. Chem. Int. Ed. 51, 1-6 (2012).
spectroscopy of interfaces
Fig.: Surface specific 2D-IR spectroscopy by sum frequency generation .
To come... For the moment please refer to this article .
We are currently running two femtosecond laser labs. Each lab features two oscillator/amplifier systems, that can be synchronized.