Determining membrane protein structures: still a challenge!

INSERM, U773, Centre de Recherche Biomédicale Bichat Beaujon CRB3, Faculté de Médecine X. Bichat, Université Paris 7, BP 416, F-75018, Paris, France.
Trends in Biochemical Sciences (Impact Factor: 13.52). 07/2007; 32(6):259-70. DOI: 10.1016/j.tibs.2007.04.001
Source: PubMed

ABSTRACT Determination of structures and dynamics events of transmembrane proteins is important for the understanding of their function. Analysis of such events requires high-resolution 3D structures of the different conformations coupled with molecular dynamics analyses describing the conformational pathways. However, the solution of 3D structures of transmembrane proteins at atomic level remains a particular challenge for structural biochemists--the need for purified and functional transmembrane proteins causes a 'bottleneck'. There are various ways to obtain 3D structures: X-ray diffraction, electron microscopy, NMR and modelling; these methods are not used exclusively of each other, and the chosen combination depends on several criteria. Progress in this field will improve knowledge of ligand-induced activation and inhibition of membrane proteins in addition to aiding the design of membrane-protein-targeted drugs.

  • [Show abstract] [Hide abstract]
    ABSTRACT: The purple membrane of Halobacterium Salinarum carries out a protein, bacteriorhodopsin (bR), which is a model for structure-function studies of membrane proteins. The heterologous expression of integral membrane proteins (IMPS) is difficult. In this study, we reported the heterologous overexpression of bacterio-opsin (bO) in Escherichia coli BL21 (DE3). Bacterio-opsin expression is facilitated by using mistic, a membrane protein from Bacillus subtilis in E. coli BL21 (DE3) membranes. The optimized bO gene was cloned in fusion to the C-terminus of mistic in pET 30a (+) and contains an oct-histidine in C-terminal to facilitate purification. Different medium, temperature, and induction time were used to optimize protein overexpression. The highest expression was obtained from the Terrific Broth (TB) medium at 18 °C with an IPTG concentration of 0.1 mM. The final purified bR was 192 ± 1 mg/L which has an important value for the production of membrane proteins in E. coli.
    Applied Biochemistry and Biotechnology 08/2014; 174(4). DOI:10.1007/s12010-014-1137-2 · 1.69 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Detergents are widely used for membrane protein research; however, membrane proteins encapsulated in micelles formed by conventional detergents tend to undergo structural degradation, necessitating the development of new agents with enhanced efficacy. Here we prepared several hydrophobic variants of ganglio-tripod amphiphiles (TPAs) derived from previously reported TPAs and evaluated for a multi-subunit, pigment protein superassembly. In this study, TPA-16 was found to be most efficient in protein solubilization while TPA-15 proved most favourable in long-term protein stability. The current study combined with previous TPA studies enabled us to elaborate on a few detergent structure-property relationships that could provide useful guidelines for novel amphiphile design.
    Organic & Biomolecular Chemistry 08/2014; 12(42). DOI:10.1039/C4OB01375A · 3.49 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The 3D structures of membrane proteins are typically determined without the presence of a lipid bilayer. For the purpose of studying the role of membranes on the wild type characteristics of the corresponding protein, determining the position and orientation of transmembrane proteins within a membrane environment is highly desirable. Here we report a geometry-based approach to automatically insert a membrane protein with a known 3D structure into pregenerated lipid bilayer membranes with various dimensions and lipid compositions or into a pseudomembrane. The pseudomembrane is built using the Protein Nano-Object Integrator which generates a parallelepiped of user-specified dimensions made up of pseudoatoms. The pseudomembrane allows for modeling the desolvation effects while avoiding plausible errors associated with wrongly assigned protein-lipid contacts. The method is implemented into a web server, the ProBLM server, which is freely available to the biophysical community. The web server allows the user to upload a protein coordinate file and any missing residues or heavy atoms are regenerated. ProBLM then creates a combined protein-membrane complex from the given membrane protein and bilayer lipid membrane or pseudomembrane. The user is given an option to manually refine the model by manipulating the position and orientation of the protein with respect to the membrane.
    Computational and Mathematical Methods in Medicine 07/2014; 2014:838259. DOI:10.1155/2014/838259 · 1.02 Impact Factor

Full-text (2 Sources)

Available from
May 29, 2014