title: Systematic analysis of protein-detergent complexes applying dynamic light scattering to optimize solutions for crystallization trials.
abstract: Detergents are widely used for the isolation and solubilization of membrane proteins to support crystallization and structure determination. Detergents are amphiphilic molecules that form micelles once the characteristic critical micelle concentration (CMC) is achieved and can solubilize membrane proteins by the formation of micelles around them. The results are presented of a study of micelle formation observed by in situ dynamic light-scattering (DLS) analyses performed on selected detergent solutions using a newly designed advanced hardware device. DLS was initially applied in situ to detergent samples with a total volume of approximately 2 µl. When measured with DLS, pure detergents show a monodisperse radial distribution in water at concentrations exceeding the CMC. A series of all-trans n-alkyl-β-D-maltopyranosides, from n-hexyl to n-tetradecyl, were used in the investigations. The results obtained verify that the application of DLS in situ is capable of distinguishing differences in the hydrodynamic radii of micelles formed by detergents differing in length by only a single CH2 group in their aliphatic tails. Subsequently, DLS was applied to investigate the distribution of hydrodynamic radii of membrane proteins and selected water-insoluble proteins in presence of detergent micelles. The results confirm that stable protein-detergent complexes were prepared for (i) bacteriorhodopsin and (ii) FetA in complex with a ligand as examples of transmembrane proteins. A fusion of maltose-binding protein and the Duck hepatitis B virus X protein was added to this investigation as an example of a non-membrane-associated protein with low water solubility. The increased solubility of this protein in the presence of detergent could be monitored, as well as the progress of proteolytic cleavage to separate the fusion partners. This study demonstrates the potential of in situ DLS to optimize solutions of protein-detergent complexes for crystallization applications.
authors.affiliation: Institute of Biochemistry and Molecular Biology, Laboratory for Structural Biology of Infection and Inflammation, University of Hamburg, c/o DESY, Building 22a, Notkestrasse 85, 22603 Hamburg, Germany.
authors.forename: Arne
authors.initials: A
authors.lastname: Meyer
authors.affiliation: Institute of Biochemistry and Molecular Biology, Laboratory for Structural Biology of Infection and Inflammation, University of Hamburg, c/o DESY, Building 22a, Notkestrasse 85, 22603 Hamburg, Germany.
authors.forename: Karsten
authors.initials: K
authors.lastname: Dierks
authors.affiliation: Institute of Biochemistry and Molecular Biology, Laboratory for Structural Biology of Infection and Inflammation, University of Hamburg, c/o DESY, Building 22a, Notkestrasse 85, 22603 Hamburg, Germany.
authors.forename: Rana
authors.initials: R
authors.lastname: Hussein
authors.affiliation: UMR 7242-IMPReSs Platform, ESBS, Pôle API, 300 Boulevard Sébastien Brant, CS10413, 67412 Illkirch CEDEX, France.
authors.forename: Karl
authors.initials: K
authors.lastname: Brillet
authors.affiliation: Multi User Center for Biomolecular Innovation, Department of Physics, São Paulo State University, UNESP/IBILCE, Caixa Postal 136, São José do Rio Preto-SP, 15054, Brazil.
authors.forename: Hevila
authors.initials: H
authors.lastname: Brognaro
authors.affiliation: Institute of Biochemistry and Molecular Biology, Laboratory for Structural Biology of Infection and Inflammation, University of Hamburg, c/o DESY, Building 22a, Notkestrasse 85, 22603 Hamburg, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Betzel
mesh_terms: D003460:Crystallization; D003902:Detergents / Q000737:chemistry*; D008027:Light; D058619:Maltose-Binding Proteins / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry*; D008823:Micelles; D008958:Models, Molecular; D010316:Particle Size; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D012542:Scattering, Radiation; D012995:Solubility; D012996:Solutions; D015534:Trans-Activators / Q000737:chemistry; D054334:Viral Regulatory and Accessory Proteins
chemical_list: D003902:Detergents; D058619:Maltose-Binding Proteins; D008565:Membrane Proteins; D008823:Micelles; D011993:Recombinant Fusion Proteins; D012996:Solutions; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein
keywords: dynamic light scattering; hydrodynamic radius; micelle size; n-alkyl-β-d-maltopyranosides; protein–detergent complex