Séminaire Optique/Spectro

Vendredi 3 Octobre 2014 à 11h00.

Gas Phase Peptide Crosslinking and UV Photodissociation Chromophores Generated by ETD

''Although the vast majority of biomolecules absorb UV photons at wavelengths less than 300 nm, the photo-reactive region of mass-selected peptides can be nudged above 300 nm by attaching synthetic dyes. Because backbone fragmentation is a result of energy redistribution, photodissociations using such modifications tend to closely resemble collisional processes (CID). Conversely, fragile chromophores have long been exploited by neutral molecule photochemists for selective fragmentation to create high-energy metastable species. One such chromophore, diazirine, results in carbenes via the photolytic loss of molecular nitrogen from the C–N–N ring. These carbenes are prominent for their ability to undergo interesting bond forming processes. Furthermore, this moiety can be easily incorporated into peptides as photoleucine (L*) via simple peptide synthesis or protein expression without changes to the native protein structure. Since bond forming reactions are not typically observed from direct activation of ionic biomolecules, UV photodissociation of peptidic diazirines presents many opportunities for unique mass spectroscopic analysis such as peptide stitching. Our primary goal is to use such stitching events to help describe the 3 dimensional structure of peptide ions in the gas phase. To achieve these goals, we have made simple modifications to a LTQ XL ETD linear ion trap mass spectrometer which allows for the photoactivation of ions using a 355 nm laser. This configuration allows for tandem analysis of ions via ETD, CID, and UV photodissociation. Our recent results show that the weak ion pair of [GLLLK + GL*L*L*K + H]+ can be covalently linked by treatment with UV light. Complementing the above tandem experiments, our instrument configuration also allows for UV photochemical investigations of the products of electron transfer dissociation (ETD). We have found that z-type fragment ions readily undergo photodissociation implying that a chromophore is created in the ETD process. Notably, the backbone fragmentations identified in this type of photodissociation are entirely different from CID. Our initial calculations indicate that the unpaired radical in nearly all z fragments is excitable at 355 nm, providing a provocative new chromophore for photodissociation sequencing methods.''