Enhancement of charge remote fragmentation in protonated peptides by high-energy CID MALDI-TOF-MS using “cold” matrices

Abstract

Delayed extraction matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (DE-MALDI-TOF-MS) is employed to evaluate its potential for peptide sequencing using both post-source decay (PSD) and high-energy collision-induced dissociation (CID). This work provides evidence that complete amino-acid sequences may be obtained employing a dual approach including PSD of [M + H] + ions using a “hot” matrix (α-cyano-4-hydroxycinnamic acid, CHCA), followed by high-energy CID using “cold” matrices (2,5-dihydroxybenzoic acid, DHB; 2,6-dihydroxyacetophenone/di-ammonium hydrogen citrate, DHAP/DAHC). This strategy ensures that PSD results in a rich variety of product ions derived from charge-driven processes that provide gross structural information. High-energy CID (20 keV collision energy range) of low internal energy [M + H] + ions is then employed to reveal complementary side-chain detail (i.e. Leu/Ile distinction) in a manner highly selective for charge remote fragmentation (CRF), because PSD is largely reduced. As expected from the known behaviour of protonated peptides at 10 keV collision energies, charge fixation at basic sites required for CRF is more pronounced in CID than in PSD. We have obtained spectra for a synthetic peptide that approximate the results and performance of MALDI high-energy CID obtained on sector-based instrumentation (EBE-oa-TOF).