title: Identifying Ca2+-binding sites in proteins by liquid chromatography-mass spectrometry using Ca2+-directed dissociations.
abstract: Here we describe a new method to identify calcium-binding sites in proteins using high-resolution liquid chromatography-mass spectrometry in concert with calcium-directed collision-induced dissociations. Our method does not require any modifications to the liquid chromatography-mass spectrometry apparatus, uses standard digestion protocols, and can be applied to existing high-resolution MS data files. In contrast to NMR, our method is applicable to very small amounts of complex protein mixtures (femtomole level). Calcium-bound peptides can be identified using three criteria: (1) the calculated exact mass of the calcium containing peptide; (2) specific dissociations of the calcium-containing peptide from threonine and serine residues; and (3) the very similar retention times of the calcium-containing peptide and the free peptide.
authors.affiliation: From the ‡Department of Neurology, Laboratory of Neuro-Oncology, Erasmus Medical Center, P.O. Box 2040, 3000 CA, Rotterdam, The Netherlands; §Thermo Fisher Scientific/Dionex, Abberdaan 114, 1046 AA, Amsterdam, The Netherlands;
authors.forename: Azadeh
authors.initials: A
authors.lastname: Jamalian
authors.affiliation: From the ‡Department of Neurology, Laboratory of Neuro-Oncology, Erasmus Medical Center, P.O. Box 2040, 3000 CA, Rotterdam, The Netherlands; §Thermo Fisher Scientific/Dionex, Abberdaan 114, 1046 AA, Amsterdam, The Netherlands;
authors.forename: Evert-Jan
authors.initials: EJ
authors.lastname: Sneekes
authors.affiliation: ¶NMR Spectroscopy, Bijvoet Center for Biomolecular Research, P.O. Box 80.075, 3508 TB, Utrecht University, Utrecht, The Netherlands;
authors.forename: Hans
authors.initials: H
authors.lastname: Wienk
authors.affiliation: From the ‡Department of Neurology, Laboratory of Neuro-Oncology, Erasmus Medical Center, P.O. Box 2040, 3000 CA, Rotterdam, The Netherlands;
authors.forename: Lennard J M
authors.initials: LJ
authors.lastname: Dekker
authors.affiliation: ‖Theoretical Chemistry Group, University of Utrecht, 3508 TC, Utrecht, The Netherlands.
authors.forename: Paul J A
authors.initials: PJ
authors.lastname: Ruttink
authors.affiliation: §Thermo Fisher Scientific/Dionex, Abberdaan 114, 1046 AA, Amsterdam, The Netherlands;
authors.forename: Mario
authors.initials: M
authors.lastname: Ursem
authors.affiliation: From the ‡Department of Neurology, Laboratory of Neuro-Oncology, Erasmus Medical Center, P.O. Box 2040, 3000 CA, Rotterdam, The Netherlands;
authors.forename: Theo M
authors.initials: TM
authors.lastname: Luider
authors.affiliation: From the ‡Department of Neurology, Laboratory of Neuro-Oncology, Erasmus Medical Center, P.O. Box 2040, 3000 CA, Rotterdam, The Netherlands; p.burgers@erasmusmc.nl.
authors.forename: Peter C
authors.initials: PC
authors.lastname: Burgers
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000502:physiology; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism*; D002645:Chickens; D002851:Chromatography, High Pressure Liquid; D051959:ELAV Proteins / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D006801:Humans; D010449:Peptide Mapping; D010920:Placenta / Q000378:metabolism; D011225:Pre-Eclampsia / Q000473:pathology; D011247:Pregnancy; D000075383:S100 Calcium Binding Protein A6; D009418:S100 Proteins / Q000737:chemistry / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014327:Trophoblasts / Q000378:metabolism
chemical_list: D002135:Calcium-Binding Proteins; D018797:Cell Cycle Proteins; D051959:ELAV Proteins; D000075383:S100 Calcium Binding Protein A6; D009418:S100 Proteins; C055351:S100A6 protein, human; D002118:Calcium