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Poster Presentations
Day 1, June 22（Sun.）
Room P (Maesato East, Foyer, Ocean Wing)
- 1P-PM-17
A Novel Benchtop MALDI-TOF/TOF Platform for Top-Down Protein Characterization
(1BRUKERSG, 2BRUKER, 3BRUKERDAL)
oWen Donq Looi1, Sergei Dikler2, Arndt Asperger3
MALDI top-down sequencing has proven to be a powerful tool for the analysis of intact proteins and protein subunits. The unique feature of MALDI in-source decay (ISD), combined with T3-sequencing (pseudo-MS3) of ISD fragment ions give MALDI TDS the capability to sequence up to 100 amino acids residues from each terminus and to elucidate detailed modifications. This capability has been applied to analyse antibodies, nanobodies, PEGylated proteins, bacterial glycoconjugates, and small membrane proteins. In this study, we demonstrate the efficacy of a novel benchtop MALDI-TOF/TOF platform in the TDS analysis of NISTmAb and other proteins. NISTmAb was first cleaved, deglycosylated and reduced, subsequently fractionated by LC using a C4 column. The resultant NISTmAb subunits, fragments and TNF-α were prepared with SDHB matrix and analyzed on a Bruker neofleX MALDI-TOF/TOF platform. MALDI-ISD and MS/MS T3 spectra were acquired in reflector mode. Results showed that MALDI-TDS was able to achieve 92.5% sequence coverage for the light chain subunit, and 83.6% sequence coverage for the Fd fragment and revealed oxidation on the Met20 residue of TNF-α fragment. T3 sequencing was able to confirm the N- and C- terminal of the light chain remained unmodified, while verifying the pGlu modification at the Fd N-terminus.