posted on 2020-04-01, 20:44authored bySimone Nicolardi, David P. A. Kilgour, Natasja Dolezal, Jan W. Drijfhout, Manfred Wuhrer, Yuri E. M. van der Burgt
Comprehensive determination
of primary sequence and identification
of post-translational modifications (PTMs) are key elements in protein
structural analysis. Various mass spectrometry (MS) based fragmentation
techniques are powerful approaches for mapping both the amino acid
sequence and PTMs; one of these techniques is matrix-assisted laser
desorption/ionization (MALDI), combined with in-source decay (ISD)
fragmentation and Fourier-transform ion cyclotron resonance (FT-ICR)
MS. MALDI-ISD MS protein analysis involves only minimal sample preparation
and does not require spectral deconvolution. The resulting MALDI-ISD
MS data is complementary to electrospray ionization-based MS/MS sequencing
readouts, providing knowledge on the types of fragment ions is available.
In this study, we evaluate the isotopic distributions of z′ ions in protein top-down MALDI-ISD FT-ICR mass spectra and
show why these distributions can deviate from theoretical profiles
as a result of co-occurring and isomeric z and y-NH3 ions. Two synthetic peptides, containing
either normal or deuterated alanine residues, were used to confirm
the presence and unravel the identity of isomeric z and y-NH3 fragment ions (“twins”).
Furthermore, two reducing MALDI matrices, namely 1,5-diaminonaphthalene
and N-phenyl-p-phenylenediamine
were applied that yield ISD mass spectra with different fragment ion
distributions. This study demonstrates that the relative abundance
of isomeric z and y-NH3 ions requires consideration for accurate and confident assignments
of z′ ions in MALDI-ISD FT-ICR mass spectra.