Structural assignment of disialylated biantennary N-glycan isomers derivatized with 2-aminopyridine using negative-ion multistage tandem mass spectral matching

To investigate the possibility of structural assignment based on negative-ion multistage tandem mass (MSn) spectral matching, four isomers of disialylated biantennary N-glycans (alpha 2-6 and/or alpha 2-3 linked sialic acid on alpha 1-6 and alpha 1-3 antennae) derivatized with 2-aminopyridine (PA) were analyzed by employing high-performance liquid chromatography/electrospray ionization linear ion trap time-of-flight mass spectrometry (HPLC/ESI-LIT-TOMS), which uses helium gas for ion trapping and collision-induced dissociation (CID). It is shown that the MS2 spectra derived from each precursor ion [M-2H](2-) are reproducible and useful for distinguishing the four isomers. Thus, they can be assigned by negative-ion MS2 spectral matching based on correlation coefficients. In addition, MS3 spectra derived from D-type fragment ions clearly differentiate the alpha 2-3- or alpha 2-6-linked sialic acid on the alpha 1-6 antenna due to their characteristic spectral patterns. The C-4-type fragment ions, which are produced from both the alpha 1-6 and alpha 1-3 antennae, show the characteristic MS3 spectra reflecting alpha 2-3- or alpha 2-6- linkage type or a mixture of both types. Thus, the differentiation and assignment of these disialylated biantennary N-glycan isomers can also be supported with the MS3 spectra of C-4- and D-type ions. Copyright (c) 2006 John Wiley & Sons, Ltd.