PROTEIN CROSS-LINKING BY UV EXCITATION

Boston University Medical Campus, Boston MA

Investigators

Linked publications, trials & patents

Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. This project intends to generate a better understanding of the cross-linking process initiated by UV excitation and determine the efficiency of this reaction. Cross-linking was performed on different peptides by UV excitation. The resulting dimers were analyzed by using a high resolution mass spectrometer (SolariX, Bruker Daltonics, Billerica, USA). We firstly demonstrated that it is possible to induce a cross-link when peptides are exposed to such a UV-laser. Tandem mass spectrometry results (CID and ECD) showed univocally that the amino acids involved in this reaction are only the aromatic ones. To determine wether or not the cross-linking reaction is related to a radical mechanism, we have performed femtosecond laser irradiation in the presence of spin trap molecules. After irradiation;we observed the formation of covalent adducts with spin trap molecules, demonstrating the radical nature of the reaction mechanism. The specific location of the adducts on the model peptides was determined by high resolution mass spectrometry, demonstrating that radical adduction occurs only on aromatic side chains. Ms. Leo spent two months at the Resource to receive training and participate in the FTMS analyses. These results were included in her PhD thesis (Jasn 2011). A manuscript is in preparation.