Crystallization of membrane proteins: bovine rhodopsin.
The conditions for crystallization of the integral membrane protein bovine rhodopsin were determined. Crystals form over a wide range of protein concentration (0.4-15 mg ml-1) and pH (5.5-7.5) in detergent solutions of octyl-polyoxyethylene (o-POE) that contain inorganic salts as precipitants. Crystallization occurs in the detergent-rich phase under conditions of phase separation. This corroborates the results of experiments on the phase behavior of the detergent solution and on the distribution of rhodopsin between the salt-rich and detergent-rich phases formed at high salt concentrations. Crystals were needle shaped, and the best crystals were obtained by vapour diffusion, at room temperature, of a 2.5 mg ml-1 protein solution in a 1.5% (v/v) o-POE solution containing 1.5 M ammonium sulphate (pH 7.0) against unbuffered 2.85 M ammonium sulphate. The crystals thus produced had dimensions of approximately 70 microns x 70 microns x 1000 microns. These crystals were too small to allow X-ray diffraction studies of rhodopsin structure, but electron microscopic studies of negatively stained thin crystals allowed the definition of the projection belonging to the two-sided plane group p21, and showed unit cell parameters alpha (50 A), b (72 A), and gamma (90 degrees).
Yurkova
E V
EV
Shemyakin Institute of Bioorganic Chemistry, Academy of Sciences of the USSR, Moscow.
Demin
V V
VV
Abdulaev
N G
NG
D000818:Animals; D002417:Cattle; D003902:Detergents / Q000494:pharmacology; D004123:Dimethylamines / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D005227:Fatty Acids / Q000494:pharmacology; D005944:Glucosamine / Q000031:analogs & derivatives / Q000494:pharmacology; D005960:Glucosides / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008854:Microscopy, Electron; D008968:Molecular Conformation; D011092:Polyethylene Glycols; D012243:Rhodopsin / Q000737:chemistry*; D013696:Temperature
D003902:Detergents; D004123:Dimethylamines; D005227:Fatty Acids; D005960:Glucosides; C072375:n-octyl-polyoxyethylene; C018619:octyl-beta-D-glucoside; D011092:Polyethylene Glycols; C014518:dodecyldimethylamine oxide; C040358:dodecyl maltoside; C042222:nonanoyl-N-methylglucamide; D012243:Rhodopsin; D005944:Glucosamine