Improved infrared multiphoton dissociation of peptides through N-terminal phosphonite derivatization.
A strategy for improving the sequencing of peptides by infrared multiphoton dissociation (IRMPD) in a linear ion trap mass spectrometer is described. We have developed an N-terminal derivatization reagent, 4-methylphosphonophenylisothiocyanate (PPITC), which allows the attachment of an IR-chromogenic phosphonite group to the N-terminus of peptides, thus enhancing their IRMPD efficiencies. After the facile derivatization process, the PPITC-modified peptides require shorter irradiation times for efficient IRMPD and yield extensive series of y ions, including those of low m/z that are not detected upon traditional CID. The resulting IRMPD mass spectra afford more complete sequence coverage for both model peptides and tryptic peptides from cytochrome c. We compare the effectiveness of this derivatization/IRMPD approach to that of a common N-terminal sulfonation reaction that utilizes 4-sulfophenylisothiocyanate (SPITC) in conjunction with CID and IRMPD.
Vasicek
Lisa A
LA
Department of Chemistry and Biochemistry, The University of Texas at Austin, Austin, Texas 78712, USA.
Wilson
Jeffrey J
JJ
Brodbelt
Jennifer S
JS
D001557:Benzenesulfonates / Q000737:chemistry; D017879:Isothiocyanates / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods*; D063065:Organophosphonates / Q000737:chemistry; D010455:Peptides / Q000737:chemistry*; D020539:Sequence Analysis, Protein / Q000379:methods*; D013861:Thiocyanates / Q000737:chemistry*
C543135:4-methylphosphonophenylisothiocyanate; D001557:Benzenesulfonates; D017879:Isothiocyanates; D063065:Organophosphonates; D010455:Peptides; D013861:Thiocyanates; C015308:4-isothiocyanatobenzenesulfonate