Involvement of the C-terminal proline-rich motif of G protein-coupled receptor kinases in recognition of activated rhodopsin.
G protein-coupled receptor kinases (GRKs) are a family of serine/threonine kinases that phosphorylate many activated G protein-coupled receptors (GPCRs) and play an important role in GPCR desensitization. Our previous work has demonstrated that the C-terminal conserved region (CC) of GRK-2 participates in interaction with rhodopsin and that this interaction is necessary for GRK-2-mediated receptor phosphorylation (Gan, X. Q., Wang, J. Y., Yang, Q. H., Li, Z., Liu, F., Pei, G., and Li, L. (2000) J. Biol. Chem. 275, 8469-8474). In this report, we further investigated whether the CC of other GRKs had the same functions and defined the specific sequences in CC that are required for the functions. The CC regions of GRK-1, GRK-2, and GRK-5, representatives of the three subfamilies of GRKs, could bind rhodopsin in vitro and inhibit GRK-2-mediated phosphorylation of rhodopsin, but not a peptide GRK substrate. Through a series of mutagenesis analyses, a proline-rich motif in the CC was identified as the key element involved in the interaction between the CC region and rhodopsin. Point mutations of this motif not only disrupted the interaction of GRK-2 with rhodopsin but also abolished the ability of GRK-2 to phosphorylate rhodopsin. The findings that the CC region of GRKs interact only with the light-activated but not the non-activated rhodopsin and that the N-terminal domain of GRK-2 interacts with rhodopsin in a light-independent manner suggest that the CC region is responsible for the recognition of activated GPCRs in the canonical model.
Gan
Xiaoqing
X
Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China.
Ma
Zhihai
Z
Deng
Ning
N
Wang
Jiyong
J
Ding
Jianping
J
Li
Lin
L
D020816:Amino Acid Motifs; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D005136:Eye Proteins / Q000378:metabolism; D051606:G-Protein-Coupled Receptor Kinase 1; D054774:G-Protein-Coupled Receptor Kinase 5; D008027:Light; D009154:Mutation; D010766:Phosphorylation; D011392:Proline / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D012243:Rhodopsin / Q000378:metabolism*; D051552:beta-Adrenergic Receptor Kinases
D005136:Eye Proteins; D043562:Receptors, G-Protein-Coupled; D012243:Rhodopsin; D011392:Proline; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D051606:G-Protein-Coupled Receptor Kinase 1; D051552:beta-Adrenergic Receptor Kinases; D054774:G-Protein-Coupled Receptor Kinase 5