Crystals of native and modified bovine rhodopsins and their heavy atom derivatives.
Rhodopsin, the pigment protein responsible for dim-light vision, is a G protein-coupled receptor that converts light absorption into the activation of a G protein, transducin, to initiate the visual response. We have crystallised detergent-solubilised bovine rhodopsin in the native form and after chemical modifications as needles 10-40 microm in cross-section. The crystals belong to the trigonal space group P3(1), with two molecules of rhodopsin per asymmetric unit, related by a non-crystallographic 2-fold axis parallel with the crystallographic screw axis along c (needle axis). The unit cell dimensions are a=103.8 A, c=76.6 A for native rhodopsin, but vary over a wide range after heavy atom derivatisation, with a between 101.5 A and 113.9 A, and c between 76.6 A and 79.2 A. Rhodopsin molecules are packed with the bundle of transmembrane helices tilted from the c-axis by about 100 degrees . The two molecules in the asymmetric unit form contacts along the entire length of their transmembrane helices 5 in an antiparallel orientation, and they are stacked along the needle axis according to the 3-fold screw symmetry. Hence hydrophobic contacts are prominent at protein interfaces both along and normal to the needle axis. The best crystals of native rhodopsin in this crystal form diffracted X-rays from a microfocused synchrotron source to 2.55 A maximum resolution. We describe steps taken to extend the diffraction limit from about 10 A to 2.6 A.
Edwards
Patricia C
PC
Medical Research Council, Laboratory of Molecular Biology, Cambridge CB2 2QH, UK.
Li
Jade
J
Burghammer
Manfred
M
McDowell
J Hugh
JH
Villa
Claudio
C
Hargrave
Paul A
PA
Schertler
Gebhard F X
GF
D000818:Animals; D002417:Cattle; D003460:Crystallization; D018360:Crystallography, X-Ray; D003902:Detergents; D019216:Metals, Heavy / Q000737:chemistry* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012160:Retina / Q000737:chemistry / Q000378:metabolism; D012243:Rhodopsin / Q000031:analogs & derivatives / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D013053:Spectrophotometry
D003902:Detergents; D019216:Metals, Heavy; D012243:Rhodopsin