2D IR OF UNUSUAL ISOTOPOMERS AND FOLDING

点击翻译按钮获取中文摘要

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. To obtain a better understanding of the process of protein folding, development of physical methods that measure the time dependences of structural changes in complex systems is essential. Because of the intrinsically high time resolution, multidimensional infrared spectroscopy (2D IR and 3D IR) can contribute significantly to this challenge. IR spectra of proteins and peptides are intimately connected with their complete three-dimensional structures on the length scale of chemical bonds. However, the vibrational transitions of proteins are not always spectrally resolved. In this respect, multidimensional nonlinear infrared methods can provide additional information on how the various modes are coupled. Isotopic replacements have been essential in the interpretation of vibrational spectra and their relationship to structure. Isotopomers have frequencies, force fields, and anharmonicities that are different from one another. Incorporation of isotopes in 2D IR leads to shift in frequencies into regions where their couplings can be measured, free from interference by other modes of the system. For the amide I mode, the shifts by 13C=18O and 13C=16O substitution are sufficiently large to displace the substituted amide group frequencies beyond the range of the natural distribution of frequencies found in most secondary structures. The strategy of the Resource is to insert both 13C=18O and 13C=16O labels into secondary structures at known residues. Since their isotope shifts are different, a pair of isotopic peaks is created and 2D IR spectroscopic methods can then be used to analyze the coupling between those specific pairs of molecular transitions. Applying this approach, the real time evolution of folding of the secondary structure of the Villin headpiece can be examined using 2D IR.