Expression of Functional G Protein-Coupled Receptors in Photoreceptors of TransgenicXenopus laevis†

Li Zhang, David Salom, Jianhua He, Alex Okun, Juan Ballesteros, Krzysztof Palczewski, Ning Li
2005 Biochemistry  
G protein-coupled receptors (GPCRs) constitute the largest superfamily of transmembrane signaling proteins; however, the only known GPCR crystal structure is that of rhodopsin. This disparity reflects the difficulty in generating purified GPCR samples of sufficient quantity and quality. Rhodopsin, the light receptor of retinal rod neurons, is produced in large amounts of homogeneous quality in the vertebrate retina. We used transgenic Xenopus laeVis to convert these retina rod cells into
more » ... tors to successfully produce 20 model GPCRs. The receptors accumulated in rod outer segments and were homogeneously glycosylated. Ligand and [ 35 S]GTPγS binding assays of the 5HT 1A and EDG 1 GPCRs confirmed that they were properly folded and functional. 5HT 1A R was highly purified by taking advantage of the rhodopsin C-terminal immunoaffinity tag common to all GPCR constructs. We have also developed an automated system that can generate hundreds of transgenic tadpoles per day. This expression approach could be extended to other animal model systems and become a general method for the production of large numbers of GPCRs and other membrane proteins for pharmacological and structural studies.
doi:10.1021/bi051386z pmid:16262251 fatcat:66yxikistrauzbyoojwc57j6o4