title: Mapping of unfolding states of integral helical membrane proteins by GPS-NMR and scattering techniques: TFE-induced unfolding of KcsA in DDM surfactant.
abstract: Membrane proteins are vital for biological function, and their action is governed by structural properties critically depending on their interactions with the membranes. This has motivated considerable interest in studies of membrane protein folding and unfolding. Here the structural changes induced by unfolding of an integral membrane protein, namely TFE-induced unfolding of KcsA solubilized by the n-dodecyl β-d-maltoside (DDM) surfactant is investigated by the recently introduced GPS-NMR (Global Protein folding State mapping by multivariate NMR) (Malmendal et al., PlosONE 5, e10262 (2010)) along with dynamic light scattering (DLS) and small-angle X-ray scattering (SAXS). GPS-NMR is used as a tool for fast analysis of the protein unfolding processes upon external perturbation, and DLS and SAXS are used for further structural characterization of the unfolding states. The combination allows addressing detergent properties and protein conformations at the same time. The mapping of the states reveals that KcsA undergoes a series of rearrangements which include expansion of the tetramer in several steps followed by dissociation into monomers at 29% TFE. Supplementary studies of DDM and TFE in the absence of KcsA suggest that the disintegration of the tetramer at 29% TFE is caused by TFE dissolving the surrounding DDM rim. Above 34% TFE, KcsA collapses to a new structure that is fully formed at 44% TFE.
authors.affiliation: Department of Chemistry, Aarhus University, Denmark.
authors.forename: Antonello
authors.initials: A
authors.lastname: Calcutta
authors.forename: Christian M
authors.initials: CM
authors.lastname: Jessen
authors.forename: Manja Annette
authors.initials: MA
authors.lastname: Behrens
authors.forename: Cristiano L P
authors.initials: CL
authors.lastname: Oliveira
authors.forename: Maria Lourdes
authors.initials: ML
authors.lastname: Renart
authors.forename: José M
authors.initials: JM
authors.lastname: González-Ros
authors.forename: Daniel E
authors.initials: DE
authors.lastname: Otzen
authors.forename: Jan Skov
authors.initials: JS
authors.lastname: Pedersen
authors.forename: Anders
authors.initials: A
authors.lastname: Malmendal
authors.forename: Niels Chr
authors.initials: NC
authors.lastname: Nielsen
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D005960:Glucosides / Q000737:chemistry; D008027:Light; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008823:Micelles; D008956:Models, Chemical; D008958:Models, Molecular; D008968:Molecular Conformation; D015221:Potassium Channels / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012542:Scattering, Radiation; D053838:Scattering, Small Angle; D013501:Surface-Active Agents / Q000737:chemistry; D014965:X-Rays
chemical_list: D001426:Bacterial Proteins; D005960:Glucosides; D008823:Micelles; D015221:Potassium Channels; D013501:Surface-Active Agents; C107790:prokaryotic potassium channel; C040358:dodecyl maltoside