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J Am Soc Mass Spectrom. 2018 Jun;29(6):1221-1229. doi: 10.1007/s13361-018-1897-y. Epub 2018 Feb 20.

Top Down Tandem Mass Spectrometric Analysis of a Chemically Modified Rough-Type Lipopolysaccharide Vaccine Candidate.

Author information

1
School of Medicine, University of Maryland, Baltimore, MD, 21201, USA.
2
National High Magnetic Field Laboratory, Florida State University, Tallahassee, FL, 32310, USA.
3
Department of Microbial Pathogenesis, School of Dentistry, University of Maryland, Baltimore, MD, 21201, USA.
4
Chemistry and Forensics, School of Science and Technology, Nottingham Trent University, Nottingham, NG11 8NS, UK.
5
Center for Vaccine Development, School of Medicine, University of Maryland, Baltimore, MD, 21201, USA.
6
Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, Pharmacy Hall North Room 623, 20 N. Pine St, Baltimore, MD, 21201, USA. dgoodlett@rx.umaryland.edu.

Abstract

Recent advances in lipopolysaccharide (LPS) biology have led to its use in drug discovery pipelines, including vaccine and vaccine adjuvant discovery. Desirable characteristics for LPS vaccine candidates include both the ability to produce a specific antibody titer in patients and a minimal host inflammatory response directed by the innate immune system. However, in-depth chemical characterization of most LPS extracts has not been performed; hence, biological activities of these extracts are unpredictable. Additionally, the most widely adopted workflow for LPS structure elucidation includes nonspecific chemical decomposition steps before analyses, making structures inferred and not necessarily biologically relevant. In this work, several different mass spectrometry workflows that have not been previously explored were employed to show proof-of-principle for top down LPS primary structure elucidation, specifically for a rough-type mutant (J5) E. coli-derived LPS component of a vaccine candidate. First, ion mobility filtered precursor ions were subjected to collision induced dissociation (CID) to define differences in native J5 LPS v. chemically detoxified J5 LPS (dLPS). Next, ultra-high mass resolving power, accurate mass spectrometry was employed for unequivocal precursor and product ion empirical formulae generation. Finally, MS3 analyses in an ion trap instrument showed that previous knowledge about dissociation of LPS components can be used to reconstruct and sequence LPS in a top down fashion. A structural rationale is also explained for differential inflammatory dose-response curves, in vitro, when HEK-Blue hTLR4 cells were administered increasing concentrations of native J5 LPS v. dLPS, which will be useful in future drug discovery efforts. Graphical Abstract ᅟ.

KEYWORDS:

Collision induced dissociation; FT-ICR; Lipid A; Lipopolysaccharide; Oligosaccharide; Top down; Vaccine

PMID:
29464544
DOI:
10.1007/s13361-018-1897-y

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