Elucidating protein structure
In addition, various theoretical approaches are also being utilized with empirically developed parameters to try to get insight into the folding mechanisms.
UV-Visible absorption spectroscopy was the first technique used to monitor the UV absorption of the peptide backbone.
In this case the vibrational modes observed are particular vibrations whose motions couple to the driven electronic motion occurring in the electronic transition.
The vibrational modes enhanced are those localized in the chromophoric segments. This results in a crucial ultra-high resonance Raman selectivity and sensitivity that makes it a very powerful technique for studying macromolecules; instead of all the sample vibrations contributing with comparable intensities, only a small subset of resonance Raman enhanced vibrations localized around the chromophoric group dominate the spectra.
UVRR excitation of Mb at 415 nm in the strong heme Soret absorption band, results in an intense UVRR spectra which contains only the in-plane heme ring vibrations.
Thus, tuning the UVRR excitation wavelengths, allows the probing of different chromophoric segments of a macromolecule.
Raman spectroscopy is an inelastic light scattering phenomenon where the incident electromagnetic field interacts with a molecule such that there is an exchange of a quantum of vibrational energy between the two, resulting in a vibrational frequency difference between the incident and scattered light.The resulting UVRR structural insight is impressive in that it allows differentiation of, for example, different α-helix-like states that enable differentiating π- and 3(10)- states from pure α-helices.