title: Defining thermostability of membrane proteins by western blotting.
abstract: Membrane proteins are relatively challenging targets for structural and other biophysical studies. Insufficient expression in various expression systems, inherent flexibility, and instability in the detergents that are required for membrane extraction are the main reasons for this limited success. Therefore, identification of suitable conditions and membrane protein variants that can help stabilize functional protein for extended periods of time is critical for structural studies. Here, we describe a western blot-based assay that simplifies identification of thermostabilizing conditions for membrane proteins. We show successful testing of a variety of parameters such as additive lipids, ligands and detergents.
authors.affiliation: Faculty of Biochemistry and Molecular Medicine & Biocenter Oulu, University of Oulu, Oulu 90014, Finland.
authors.forename: Y
authors.initials: Y
authors.lastname: Ashok
authors.affiliation: Faculty of Biochemistry and Molecular Medicine & Biocenter Oulu, University of Oulu, Oulu 90014, Finland.
authors.forename: R
authors.initials: R
authors.lastname: Nanekar
authors.affiliation: Faculty of Biochemistry and Molecular Medicine & Biocenter Oulu, University of Oulu, Oulu 90014, Finland Present address: Center for Proteomic Chemistry, Novartis Institute of Biomedical Research, Basel, Switzerland veli-pekka.jaakola@oulu.fi.
authors.forename: V-P
authors.initials: VP
authors.lastname: Jaakola
mesh_terms: D015153:Blotting, Western / Q000379:methods*; D003902:Detergents; D008055:Lipids; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D018583:Pliability; D015202:Protein Engineering; D055550:Protein Stability
chemical_list: D003902:Detergents; D008055:Lipids; D008565:Membrane Proteins
keywords: adenosine A2a receptor; membrane protein; thermostabilization; western blotting