Iodo-Labeling of Peptides for Quantitative MALDI MS Analysis─Screening for Bacteria-Binding Peptides from a Glycine-Zipper Library.

Abstract

Matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) is widely valued for its speed and sensitivity in biomolecular analysis, yet the inherently nonquantitative nature hampers its use in many applications including high-throughput screening. Here, we introduce an iodo-based labeling strategy that enables accurate quantification of peptides and peptide libraries using high-resolution MALDI FT-ICR MS. The peptides are coupled at the N-terminus with benzoic acid (BA) or 4-iodobenzoic acid (IBA) to generate the analyte and its internal standard, respectively, differing only by a single iodine substitution. This new labeling strategy was first validated using a simple four-peptide mixture, and subsequently applied to quantitatively evaluate glycine-zipper peptide libraries containing up to 125 members for the discovery of bacterial-binding peptides. Screening of these libraries against Gram-negative Escherichia coli and Gram-positive Bacillus subtilis revealed peptides with strong and selective interactions with the bacteria. This universally applicable, cost-effective, and straightforward approach for peptide quantification significantly enhances the reliability and accuracy of high-throughput peptide screening via MALDI FT-ICR MS.