Probing Protein Structure with Ultraviolet Photodissociation
In top-down proteomics studies proteins are characterized by iterative fragmentation and analysis via mass spectrometry. Different fragmentation techniques have their own strengths and weaknesses, and ultraviolet photodissociation (UVPD) offers the benefit of relatively consistent fragmentation along the protein backbone. In our most recent publication, we show that the technique also can distinguish subtle differences in the protein structure .
We used ion-mobility mass-spectrometry to resolve two distinct conformations of the 11+ charge state of the protein ubiquitin. UVPD fragmentation of those conformers, which are identical in their chemical composition and charge, reveals significantly different fragmentation patterns, which arise mostly from cleavage around a single residue, proline-19. Using both spectral data and molecular dynamics simulations, we could attribute these fragmentation differences, to cis-trans isomerization of the peptide bond N-terminal to that residue.
Our data provides first evidence for UVPD being sensitive to the higher order structure of proteins. Furthermore, conformer-selective UVPD can potentially serve as tool for the structural analysis of proteins in the gas phase.