Efficient coupling of transducin to monomeric rhodopsin in a phospholipid bilayer.
G protein-coupled receptors (GPCRs) are seven transmembrane domain proteins that transduce extracellular signals across the plasma membrane and couple to the heterotrimeric family of G proteins. Like most intrinsic membrane proteins, GPCRs are capable of oligomerization, the function of which has only been established for a few different receptor systems. One challenge in understanding the function of oligomers relates to the inability to separate monomeric and oligomeric receptor complexes in membrane environments. Here we report the reconstitution of bovine rhodopsin, a GPCR expressed in the retina, into an apolipoprotein A-I phospholipid particle, derived from high density lipoprotein (HDL). We demonstrate that rhodopsin, when incorporated into these 10 nm reconstituted HDL (rHDL) particles, is monomeric and functional. Rhodopsin.rHDL maintains the appropriate spectral properties with respect to photoactivation and formation of the active form, metarhodopsin II. Additionally, the kinetics of metarhodopsin II decay is similar between rhodopsin in native membranes and rhodopsin in rHDL particles. Photoactivation of monomeric rhodopsin.rHDL also results in the rapid activation of transducin, at a rate that is comparable with that found in native rod outer segments and 20-fold faster than rhodopsin in detergent micelles. These data suggest that monomeric rhodopsin is the minimal functional unit in G protein activation and that oligomerization is not absolutely required for this process.
Whorton
Matthew R
MR
Department of Pharmacology, University of Michigan Medical School, Ann Arbor, Michigan 48109-0632, USA.
Jastrzebska
Beata
B
Park
Paul S-H
PS
Fotiadis
Dimitrios
D
Engel
Andreas
A
Palczewski
Krzysztof
K
Sunahara
Roger K
RK
D000818:Animals; D002417:Cattle; D006801:Humans; D008051:Lipid Bilayers; D046529:Microscopy, Electron, Transmission; D008958:Models, Molecular; D010743:Phospholipids / Q000378:metabolism*; D011485:Protein Binding; D012243:Rhodopsin / Q000737:chemistry / Q000378:metabolism*; D015293:Transducin / Q000378:metabolism*
D008051:Lipid Bilayers; D010743:Phospholipids; D012243:Rhodopsin; D015293:Transducin