title: Pre-equilibration kinetic size-exclusion chromatography with mass spectrometry detection (peKSEC-MS) for label-free solution-based kinetic analysis of protein-small molecule interactions.
abstract: Here we introduce pre-equilibration kinetic size-exclusion chromatography with mass-spectrometry detection (peKSEC-MS), which is a label-free solution-based kinetic approach for characterizing non-covalent protein-small molecule interactions. In this method, a protein and a small molecule are mixed outside the column and incubated to approach equilibrium. The equilibrium mixture is then introduced into the SEC column to initiate the dissociation process by separating small molecules from the complex inside the column. A numerical model of a 1-dimensional separation was constructed to simulate mass chromatograms of the small molecule for varying rate constants of binding.
authors.affiliation: Department of Chemistry and Centre for Research on Biomolecular Interactions, York University, Toronto, Ontario M3J 1P3, Canada. skrylov@yorku.ca.
authors.forename: Jiayin
authors.initials: J
authors.lastname: Bao
authors.forename: Svetlana M
authors.initials: SM
authors.lastname: Krylova
authors.forename: Leonid T
authors.initials: LT
authors.lastname: Cherney
authors.forename: J C Yves
authors.initials: JC
authors.lastname: Le Blanc
authors.forename: Patrick
authors.initials: P
authors.lastname: Pribil
authors.forename: Philip E
authors.initials: PE
authors.lastname: Johnson
authors.forename: Derek J
authors.initials: DJ
authors.lastname: Wilson
authors.forename: Sergey N
authors.initials: SN
authors.lastname: Krylov
mesh_terms: D002850:Chromatography, Gel / Q000295:instrumentation / Q000379:methods*; D004867:Equipment Design; D007700:Kinetics; D013058:Mass Spectrometry / Q000295:instrumentation / Q000379:methods*; D008727:Methotrexate / Q000378:metabolism*; D008954:Models, Biological; D011485:Protein Binding; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism*
chemical_list: D013762:Tetrahydrofolate Dehydrogenase; D008727:Methotrexate