Oral Presentation 23rd Annual Lorne Proteomics Symposium 2018

Improving the MALDI fragmentation pattern of complex and intact disulphide bonds with aniline (#64)

Evelyne Maes 1 , Jolon M Dyer 1 2 3 4 , Santanu Deb-Choudhury 1 , Stefan Clerens 1 2
  1. AgResearch Limited, Lincoln, New Zealand
  2. Biomolecular Interaction Centre, University of Canterbury, Christchurch, New Zealand
  3. Riddet Institute, Massey University, Palmerston North, New Zealand
  4. Wine, Food & Molecular Biosciences, Lincoln University, Lincoln, Christchurch, New Zealand

Characterization of peptides containing intact disulphide bonds (DSB) via mass spectrometry is challenging. DSB are key post-translational modifications in proteins and highly important in the overall stabilization of protein structure. Mapping these DSB and determining the cysteine residue pairing delivers crucial information in protein characterization. Although applications to decipher DSB are not limited to antibodies (biologicals), they may be the major driver in pushing the field forward, as mapping of disulphide patterns in antibodies is crucial to determine the structural characteristics of the protein therapeutics. While most electrospray (ESI)-based applications mostly provide large-scale automated analysis, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) is applied to quickly check the cross-link presence. Our study demonstrates that the addition of aniline to alpha-cyano-4-hydroxycinnamic acid as matrix allows a more efficient detection and fragmentation of peptides containing DSB in non-reduced proteins. We report how incorporating aniline in a common MALDI matrix, alpha-cyano-4-hydroxycinnamic acid, improves both DSB detection in MS as well as DSB peptide fragmentation behaviour by MALDI-TOF-TOF compared to other additives and matrices. This improved disulphide assignment will be a significant new tool for when a simple screening to confirm the DSB existence is required.