CLONING AND EXPRESSION OF GRK5 - A MEMBER OF THE G-PROTEIN-COUPLED RECEPTOR KINASE FAMILY

Guanine nucleotide binding protein (G-protein)-coupled receptor kinase s (GRKs) specifically phosphorylate the agonist-occupied form of G-pro tein-coupled receptors such as the beta2-adrenergic receptor and rhodo psin. The best characterized members of this family include the beta-a drenergic receptor kinase (betaARK) and rhodopsin kinase. To identify additional members of the GRK family, the polymerase chain reaction wa s used to amplify human heart cDNA using degenerate oligonucleotide pr imers from highly conserved regions unique to the GRK family. Here we report the isolation of a cDNA that encodes a 590-amino acid protein k inase, termed GRK5, which has 34.8% and 47.2% amino acid identities wi th betaARK and rhodopsin kinase, respectively. Interestingly, GRK5 has an even higher homology with Drosophila GPRK-2 (71.0% identity) and t he recently identified human IT11 (69.1 % identity). Northern blot ana lysis of GRK5 with selected human tissues reveals a message of almost- equal-to 3 kilobases with highest levels in heart, placenta, lung > sk eletal muscle > brain, liver, pancreas > kidney. GRK5, overexpressed i n Sf9 insect cells using the baculovirus system, was able to phosphory late rhodopsin in a light-dependent manner. In addition, GRK5 neither contains a consensus sequence for isoprenylation like rhodopsin kinase nor is activated by G-protein betagamma subunits like betaARK1. Thus, GRK5 represents a member of the GRK family that likely has a unique p hysiological role.