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Chemical cross‐linking with NHS esters: a systematic study on amino acid reactivities
Author(s) -
Mädler Stefanie,
Bich Claudia,
Touboul David,
Zenobi Renato
Publication year - 2009
Publication title -
journal of mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 1076-5174
DOI - 10.1002/jms.1544
Subject(s) - chemistry , amino acid , reactivity (psychology) , succinimide , serine , tyrosine , nucleophile , threonine , lysine , tandem mass spectrometry , primary (astronomy) , stereochemistry , mass spectrometry , combinatorial chemistry , organic chemistry , biochemistry , chromatography , enzyme , catalysis , medicine , alternative medicine , pathology , physics , astronomy
Structure elucidation of tertiary or quaternary protein structures by chemical cross‐linking and mass spectrometry (MS) has recently gained importance. To locate the cross‐linker modification, dedicated software is applied to analyze the mass or tandem mass spectra (MS/MS). Such software requires information on target amino acids to limit the data analysis time. The most commonly used homobifunctional N‐hydroxy succinimide (NHS) esters are often described as reactive exclusively towards primary amines, although side reactions with tyrosine and serine have been reported. Our goal was to systematically study the reactivity of NHS esters and derive some general rules for their attack of nucleophilic amino acid side chains in peptides. We therefore studied the cross‐linking reactions of synthesized and commercial model peptides with disuccinimidyl suberate (DSS). The first reaction site in all cases was expectedly the α‐NH 2 ‐group of the N ‐terminus or the ε‐NH 2 ‐group of lysine. As soon as additional cross‐linkers were attached or loops were formed, other amino acids were also involved in the reaction. In addition to the primary amino groups, serine, threonine and tyrosine showed significant reactivity due to the effect of neighboring amino acids by intermediate or permanent Type‐1 cross‐link formation. The reactivity is highly dependent on the pH and on adjacent amino acids. Copyright © 2009 John Wiley & Sons, Ltd.