Determination of the molecular mass and dimensions of membrane proteins by size exclusion chromatography.

TitleDetermination of the molecular mass and dimensions of membrane proteins by size exclusion chromatography.
Publication TypeJournal Article
Year of Publication2008
AuthorsKunji, ERS, Harding, M, P Butler, JG, Akamine, P
JournalMethods
Volume46
Issue2
Pagination62-72
Date Published2008 Oct
ISSN1095-9130
KeywordsAnimals, Cattle, Chemical Phenomena, Chromatography, Gel, Detergents, Glucosides, Hydrophobic and Hydrophilic Interactions, Membrane Proteins, Micelles, Mitochondrial ADP, ATP Translocases, Models, Molecular, Molecular Weight, Saccharomyces cerevisiae Proteins
Abstract

Size exclusion chromatography is an established technique for the determination of hydrodynamic volumes of proteins or protein complexes. When applied to membrane proteins, the contribution of the detergent micelle, which is required to keep the protein soluble in the aqueous phase, needs to be determined to obtain accurate measurements for the protein. In a detergent series, in which the detergents differ only by the length of the alkyl chain, the contribution of the detergent micelle to the hydrodynamic volume is variable, whereas the contribution of the protein is constant. By using this approach, several parameters of membrane proteins can be estimated by extrapolation, such as the radius at the midpoint of the membrane, the average radius, the Stokes radius, and the excluded volume. The molecular mass of the protein can be determined by two independent measurements that arise from the behaviour of the free detergent micelle and protein-detergent micelle during size exclusion chromatography and the determination of the detergent-protein ratio. Determining the dimensions of protein-detergent micelles may facilitate membrane protein purification and crystallization by defining the accessibility of the protein surface.

DOI10.1016/j.ymeth.2008.10.020
Alternate JournalMethods
Citation Key10.1016/j.ymeth.2008.10.020
PubMed ID18952172
Grant ListMC_U105663139 / / Medical Research Council / United Kingdom