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Quantitative determination of lysozyme‐ligand binding in the solution and gas phases by electrospray ionisation mass spectrometry
Author(s) -
Veros Christopher T.,
Oldham Neil J.
Publication year - 2007
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.3232
Subject(s) - chemistry , lysozyme , infrared multiphoton dissociation , dissociation (chemistry) , electrospray ionization , mass spectrometry , electrospray , chromatography , analytical chemistry (journal) , collision induced dissociation , crystallography , tandem mass spectrometry , biochemistry
Affinity constants for the binding of a range of substrate and non‐substrate oligosaccharides to hen egg white lysozyme were determined by direct observation of the protein.ligand complexes using electrospray ionisation mass spectrometry (ESI‐MS) with a chip‐based nano‐ESI source. The values obtained for a series of β ‐1,4‐ N ‐acetylglucosamine oligomers (NAG n ) were found to be in good agreement with those determined by fluorescence measurement. Oligomers of α ‐1,4‐glucose (Glc n ), which are believed to bind to lysozyme non‐specifically, exhibited a 10 6 ‐ to 10 8 ‐fold lower affinity for the enzyme. Lysozyme.NAG n complexes displayed an increase in K a from n = 2 to n = 4, but then reached a plateau. In contrast non‐specific lysozyme.Glc n complexes showed no such trend. Determination of gas‐phase complex stability was achieved by quantitative collision‐induced dissociation (CID) and infrared multiphoton dissociation (IRMPD) measurements. The collision energy (Ec 50 ) or laser power (IRMPD 50 ) required to dissociate precursor ions to 50% of their original intensity was determined for lysozyme.NAG n and Glc n complexes using the [M+8H] 8+ charge state. An excellent correlation between trends in K a and gas‐phase stability was seen for NAG n oligomers bound to lysozyme, whereas no such relationship was observed with the non‐specific, weaker lysozyme.Glc n complexes. These results illustrate that ESI‐MS can be used to quantify the interactions between lysozyme and oligosaccharides in both the solution and gas phase and that measurement of gas‐phase complex stability by CID or IRMPD can provide information about specific solution binding events. Copyright © 2007 John Wiley & Sons, Ltd.