Premium
Comparison of negative and positive ion electrospray ionization mass spectra of calmodulin and its complex with trifluoperazine
Author(s) -
Watt Stephen J.,
Oakley Aaron,
Sheil Margaret M.,
Beck Jennifer L.
Publication year - 2005
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.2039
Subject(s) - chemistry , calmodulin , trifluoperazine , electrospray ionization , mass spectrum , ion , mass spectrometry , conformational change , analytical chemistry (journal) , crystallography , calcium , stereochemistry , chromatography , organic chemistry
Abstract The protein calmodulin (apoCaM) undergoes a conformational change when it binds calcium. This structure of the protein (Ca 4 CaM) is a dumbbell‐shaped molecule that undergoes a further profound conformational change on binding of the antipsychotic drug trifluoperazine (TFP). Experimental conditions were developed to prepare samples of apoCaM, Ca 4 CaM and Ca 4 CaM/TFP that were substantially free of sodium. The effects of the conformational changes of calmodulin on the charge‐state distributions observed in positive ion and negative ion electrospray ionization (ESI) mass spectra were examined. Conversion of apoCaM into Ca 4 CaM was concomitant with a change in the negative ion ESI mass spectrum whereby the 16− ion was the most abundant ion observed for the apo form and the 8− ion was the most abundant for the complex. In contrast, in the positive ion ESI mass spectra of apoCaM and Ca 4 CaM, the most abundant species in each case was the 8+ ion. When a complex of Ca 4 CaM with TFP was prepared, the most abundant species was the 5+ ion. This is consistent with a conformational change of Ca 4 CaM that rendered some basic sites inaccessible to ionization in the ESI process. Using the same Ca 4 CaM/TFP mixture, no complex with TFP was observed in negative ion ESI mass spectra. These observations are discussed in the context of the structural changes that are known to occur in calmodulin, and suggestions are made to explain the apparently conflicting data. The results reported here reflect on the validity of using differences in charge‐state distributions observed in ESI mass spectra to assess conformational changes in proteins. Copyright © 2005 John Wiley & Sons, Ltd.