Characterization of Native Protein Complexes Using Ultraviolet Photodissociation Mass Spectrometry

Abstract

Ultraviolet photodissociation (UVPD) mass spectrometry (MS) was used to characterize the sequences of proteins in native protein-ligand and protein-protein complexes and to provide auxiliary information about the binding sites of the ligands and protein-protein interfaces. UVPD outperformed collisional induced dissociation (CID) and higher-energy collisional dissociation (HCD) in terms of yielding the most comprehensive diagnostic primary sequence information of the proteins in the complexes. UVPD also generated non-covalent fragment ions containing a portion of the protein still bound to the ligand which revealed some insight into the nature of the binding sites of myoglobin/heme, eIF4E/m7GTP, and as well as human peptidyl-prolyl cis-trans isomerase Pin1 in complex with the peptide derived from the C-terminal domain of RNA polymerase II. Non-covalently bound protein-protein fragment ions from oligomeric beta-lactoglobulin dimers and hexameric insulin complexes were also produced upon UVPD, providing some illumination of tertiary and quaternary protein structural features.