Peptide-Based Drug Design

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Sequence Analysis of Antimicrobial Peptides by Tandem
Mass Spectrometry
Christin Stegemann and Ralf Hoffmann
Summary
The emergence of new diseases as well as the increasing resistance of bacteria against
antibiotics over the past decades has become a growing threat for humans. This has driven
a sustained search for new agents that possess antibacterial activities against bacteria being
resistant against conventional antibiotics and prompted an interest in short to mediumsized
peptides called antimicrobial peptides (AMPs). Such peptides were isolated from
different species, including mammals, insects, birds, fish, and plants. As these peptides
circulate only at low concentrations in body fluids, a multidimensional purification strategy
is obligatory to obtain pure peptides. The resulting low peptide amounts require highly
sensitive analytical techniques to sequence the peptides, such as Edman degradation or mass
spectrometry. Here we describe the protocols used routinely in our laboratory to identify
peptides with antimicrobial activities by mass spectrometry including de novo sequence
analysis.
Key Words: Antibacterial peptide; collision-induced dissociation (CID); electrospray
ionization (ESI); matrix-assisted laser desorption/ionization (MALDI); peptide derivatization.
1. Introduction
Despite remarkable advances in antibiotic therapy over the last century, the
incidence of serious bacterial and fungal infections in humans is increasing
due to the rapid emergence of drug-resistant Gram-positive and Gram-negative
bacterial strains (1–3). Thus one of the most urgent topics is the search for
new compound classes which exert their antimicrobial activity by molecular
mechanisms different from the currently used antibiotics. One of the most
From: Methods in Molecular Biology, vol. 494: Peptide-Based Drug Design
Edited by: L. Otvos, DOI: 10.1007/978-1-59745-419-3 3, © Humana Press, New York, NY
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