Journal
ANALYTICAL CHEMISTRY
Volume 82, Issue 6, Pages 2170-2174Publisher
AMER CHEMICAL SOC
DOI: 10.1021/ac902633d
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Funding
- Natural Sciences and Engineering Research Council of Canada
- Alberta Ingenuity Centre for Carbohydrate Science
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The application of the direct electrospray ionization mass spectrometry (ES-MS) assay for quantifying the stoichiometry and absolute affinity of protein-metal ion binding in vitro is described. Control ES-MS experiments performed on solutions containing calcium chloride or calcium acetate and a pair of proteins that do not bind calcium ions in solution revealed that the nonspecific association of metal ions to proteins during ES is a random process, independent of protein size and structure. These results establish the reliability of the reference protein method for quantitatively correcting ES mass spectra for the occurrence of nonspecific metal ion binding to proteins during ES-MS analysis. To demonstrate the utility of the direct ES-MS assay, when carried out using the reference protein method, the calcium binding stoichiometry of bovine alpha-lactalbumin and the calcium ion affinity of bovine beta-lactoglobulin were established.
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