Charge derivatization by 4-sulfophenyl isothiocyanate enhances peptide sequencing by post-source decay matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

• Published in: Journal of mass spectrometry. Vol. 38; no. 4; pp. 373 - 377
• Main Authors: Marekov, Lyuben N., Steinert, Peter M.
• Format: Journal Article
• Language: English
• Published: Chichester, UK John Wiley & Sons, Ltd 01.04.2003; Wiley
• Subjects: Amino Acid Sequence; Aminoacids, peptides. Hormones. Neuropeptides; Analytical, structural and metabolic biochemistry; Benzenesulfonates - chemistry; Biological and medical sciences; Calcium-Binding Proteins - chemistry; Calcium-Binding Proteins - metabolism; charge derivatization; Fundamental and applied biological sciences. Psychology; Humans; Isothiocyanates - chemistry; matrix-assisted laser desorption/ionization time-of-flight mass spectrometry; Molecular Sequence Data; Peptide Fragments - chemistry; peptides; post-source decay; Proteins; Sequence Analysis, Protein; sequencing; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Transglutaminases - chemistry; Transglutaminases - metabolism; Trypsin - metabolism; charge derivatization; peptides; Matrix assisted laser desorption ionization; Derivatization; Sulfonic acid; Protein; Binding protein; Time of flight method; Organic isothiocyanate; matrix-assisted laser desorption/ionization time-of-flight mass spectrometry; post-source decay; Mass spectrometry; Sequencing; Calmodulin; Structural analysis
• ISSN: 1076-5174; 1096-9888
• DOI: 10.1002/jms.448

High‐sensitivity, rapid identification of proteins in proteomic studies normally uses a combination of one‐ or two‐dimensional electrophoresis together with mass spectrometry. The simplicity and sensitivity of matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOFMS) have increased its application in recent years. The most common method of ‘peptide fingerprinting’ often may not provide robust identification. Normally additional sequence information by post‐source decay (PSD) MALDI‐TOFMS provides additional constraints for database searches to achieve highly confident results. Here we describe a derivatization procedure to facilitate the acquisition of such sequence information. Peptide digests from a skin‐expressed protein were modified with 4‐sulfophenyl isothiocyanate. The resulting peptides carry a fixed negative charge at the N‐terminal end and the resulting PSD spectrum is dominated by C‐terminal y‐type ions. The sequence information in most cases can be obtained manually or with simple programming tools. Methods of optimizing the procedure and increasing the sensitivity are discussed. Copyright © 2003 John Wiley & Sons, Ltd.