skip to main content

Title: Systematic analysis of protein–detergent complexes applying dynamic light scattering to optimize solutions for crystallization trials

Application of in situ dynamic light scattering to solutions of protein–detergent complexes permits characterization of these complexes in samples as small as 2 µl in volume. 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-transn-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 CH{sub 2} group in their aliphatic tails. Subsequently, DLS was applied to investigate the distribution of hydrodynamic radiimore » 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.« less
Authors:
 [1] ;  [1] ;  [2] ;  [1] ;  [3] ;  [4] ;  [1]
  1. University of Hamburg, c/o DESY, Building 22a, Notkestrasse 85, 22603 Hamburg (Germany)
  2. (Germany)
  3. ESBS, Pôle API, 300 Boulevard Sébastien Brant, CS10413, 67412 Illkirch CEDEX (France)
  4. São Paulo State University, UNESP/IBILCE, Caixa Postal 136, São José do Rio Preto-SP, 15054 (Brazil)
Publication Date:
OSTI Identifier:
22375711
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta crystallographica. Section F, Structural biology communications; Journal Volume: 71; Journal Issue: Pt 1; Other Information: PMCID: PMC4304753; PMID: 25615974; PUBLISHER-ID: nj5204; PUBLISHER-ID: S2053230X14027149; OAI: oai:pubmedcentral.nih.gov:4304753; Copyright (c) Meyer et al. 2015; This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CLEAVAGE; CRYSTALLIZATION; EQUIPMENT; LENGTH; LIGANDS; LIGHT SCATTERING; MATHEMATICAL SOLUTIONS; MEMBRANES; MOLECULES; POTENTIALS; SOLUBILITY; SOLUTIONS; SPATIAL DISTRIBUTION; WATER