Use of octyl beta-thioglucopyranoside in two-dimensional crystallization of membrane proteins.
A great interest exists in producing and/or improving two-dimensional (2D) crystals of membrane proteins amenable to structural analysis by electron crystallography. Here we report on the use of the detergent n-octyl beta-d-thioglucopyranoside in 2D crystallization trials of membrane proteins with radically different structures including FhuA from the outer membrane of Escherichia coli, light-harvesting complex II from Rubrivivax gelatinosus, and Photosystem I from cyanobacterium Synechococcus sp. We have analyzed by electron microscopy the structures reconstituted after detergent removal from lipid-detergent or lipid-protein-detergent micellar solutions containing either only n-octyl beta-d-thioglucopyranoside or n-octyl beta-d-thioglucopyranoside in combination with other detergents commonly used in membrane protein biochemistry. This allowed the definition of experimental conditions in which the use of n-octyl beta-d-thioglucopyranoside could induce a considerable increase in the size of reconstituted membrane structures, up to several micrometers. An other important feature was that, in addition to reconstitution of membrane proteins into large bilayered structures, this thioglycosylated detergent also was revealed to be efficient in crystallization trials, allowing the proteins to be analyzed in large coherent two-dimensional arrays. Thus, inclusion of n-octyl beta-d-thioglucopyranoside in 2D crystallization trials appears to be a promising method for the production of large and coherent 2D crystals that will be valuable for structural analysis by electron crystallography and atomic force microscopy.
Chami
M
M
Section de Recherche, Institut Curie, UMR-CNRS 168 et LRC-CEA 8, 11 rue Pierre et Marie Curie, 75231 Paris, France.
Pehau-Arnaudet
G
G
Lambert
O
O
Ranck
J L
JL
Lèvy
D
D
Rigaud
J L
JL
D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D003460:Crystallization; D000458:Cyanobacteria; D003902:Detergents / Q000378:metabolism*; D004926:Escherichia coli; D029968:Escherichia coli Proteins; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D008823:Micelles; D008854:Microscopy, Electron; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D020560:Proteobacteria; D011991:Receptors, Virus / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D013863:Thioglucosides / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D003902:Detergents; D029968:Escherichia coli Proteins; C050900:FhuA protein, E coli; D008081:Liposomes; D008565:Membrane Proteins; D008823:Micelles; D045322:Photosynthetic Reaction Center Complex Proteins; D011991:Receptors, Virus; D013863:Thioglucosides; C042989:n-octyl-beta-D-thioglucopyranoside