title: Determination of peptide topology through time-resolved double-resonance under electron capture dissociation conditions.
abstract: Characterizing the conformation of biomolecules by mass spectrometry still represents a challenge. With their knotted structure involving a N-terminal macrolactam ring where the C-terminal tail of the peptide is threaded and sterically trapped, lasso peptides constitute an attractive model for developing methods for characterizing gas-phase conformation, through comparison with their unknotted topoisomers. Here, the kinetics of electron capture dissociation (ECD) of a lasso peptide, capistruin, was investigated by electrospray ionization-Fourier transform ion cyclotron resonance mass spectrometry and compared to that of its branched-cyclic topoisomer, lactam-capistruin. Both peptides produced rather similar ECD spectra but showed different extent of H(•) transfer from c(i)' to z(j)(•) ions. Time-resolved double-resonance experiments under ECD conditions were performed to measure the formation rate constants of typical product ions. Such experiments showed that certain product ions, in particular those related to H(•) transfer, proceeded through long-lived complexes for capistruin, while fast dissociation processes predominated for lactam-capistruin. The formation rate constants of specific ECD product ions enabled a clear differentiation of the lasso and branched-cyclic topoisomers. These results indicate that the formation kinetics of ECD product ions constitute a new way to explore the conformation of biomolecules and distinguish between topoisomers and, more generally, conformers.
authors.affiliation: National Museum of Natural History, Communication Molecules and Adaptation of Microorganisms, UMR 7245 CNRS-MNHN, CP 54, 57 rue Cuvier, F-75005 Paris, France.
authors.forename: Marie
authors.initials: M
authors.lastname: Pérot-Taillandier
authors.forename: Séverine
authors.initials: S
authors.lastname: Zirah
authors.forename: Sylvie
authors.initials: S
authors.lastname: Rebuffat
authors.forename: Uwe
authors.initials: U
authors.lastname: Linne
authors.forename: Mohamed A
authors.initials: MA
authors.lastname: Marahiel
authors.forename: Richard B
authors.initials: RB
authors.lastname: Cole
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Tabet
authors.forename: Carlos
authors.initials: C
authors.lastname: Afonso
mesh_terms: D017357:Cyclotrons; D004583:Electrons; D005740:Gases; D007477:Ions; D010455:Peptides / Q000032:analysis*; D011487:Protein Conformation; D011522:Protons; D021241:Spectrometry, Mass, Electrospray Ionization; D017550:Spectroscopy, Fourier Transform Infrared; D013237:Stereoisomerism; D053719:Tandem Mass Spectrometry; D014732:Vibration
chemical_list: D005740:Gases; D007477:Ions; D010455:Peptides; D011522:Protons; C533129:capistruin