Ultraviolet photodissociation and electron transfer dissociation for peptides and oligosaccharides in quadrupole ion trap using chemical derivatization

Photodissociation methods have been explored for structural analysis of peptides and oligosaccharides. Ultraviolet photodissociation (UVPD) was applied to carboxylated derivatized peptides and reducing end derivatized oligosaccharides which offer selective dissociation and specific fragmentation pathways in comparison to CID. Upon UVPD of the modified peptides at carboxylate comprised of reduced y ions and increased immonium ions. The derivatized oligosaccharides via reductive amination and hydrazide conjugation can undergo highly efficient 355 nm UVPD and offer different fragmentation pathways. Both derivatization methods upon UVPD yielded [superscript 0,2] A-type ions, however reductive amination and hydrazide conjugation produced dominant [superscript 0,1] A and [superscript 2,4] A-type ions, respectively. Ultraviolet photodissociation at 193 nm (ArF laser, 6.4 eV / photon) has been applied to sialylated oligosaccharides and glycans which were analyzed in negative mode due to their acidic condition. Primarily, UVPD provides a greater array of fragment ions including cross-ring cleavages and dual cleavage internal ions in comparison to CID. In addition, the UVPD generates unique fragment ions which arise from site-specific cleavage of the trial substituent of the sialic acid residue. UVPD of doubly deprotonated sialylated oligosaccharides produced mostly singly deprotonated fragment ions, whereas the product ions in the CID spectra were overwhelmingly doubly charged ions, an outcome attributed to the more extensive cleavages of sialic acid residue upon UVPD. Although electron transfer dissociation (ETD) has shown superior capabilities for the characterization of post-translational modifications of peptides due to its non-eragodic property, ETD has intrinsic drawback arising from its significant dependence on the charge state of the selected precursor ion. Precursor ions in low charge states tend to undergo charge reduction, often preferentially relative to production of the informative cand z-type ions. In order to increase charge states of peptides and ETD efficiencies, peptides were derivatized at their carboxylate groups via attachment of amine with fixed charge or hydrophobic group. The carboxylate-derivatized peptides exhibited higher ETD efficiencies relative to underivatized peptides along with greater numbers of diagnostic fragment ions. The carboxylate derivatization strategy in combination with ETD for proteomics applications by the proteolytic digestion, the derivatization, and LC-MS purification was demonstrated with Cytochrome C.