Gan X Q, Wang J Y, Yang Q H, Li Z, Liu F, Pei G, Li L
Shanghai Institute of Biochemistry, Shanghai 200031, People's Republic of China.
J Biol Chem. 2000 Mar 24;275(12):8469-74. doi: 10.1074/jbc.275.12.8469.
The C-terminal domain of G protein-coupled receptor kinases (GRKs) consists of a conserved region and a variable region, and the variable region has been shown to direct the membrane translocation of cytosolic enzymes. The present work has revealed that the C-terminal domain may also be involved in kinase-receptor interaction that is primarily mediated by the conserved region. Truncation of the C-terminal domain or deletion of the conserved region in this domain of GRK2 resulted in a complete loss of its ability to phosphorylate rhodopsin and in an obvious decrease in its sensitivity to receptor-mediated phosphorylation of a peptide substrate. On the contrary, deletion of the betagamma subunit binding region in the C-terminal domain of GRK2 did not significantly alter the ability of the enzyme to phosphorylate rhodopsin. In addition, the recombinant proteins that represent the C-terminal domain and the conserved region of GRK2 could inhibit GRK2-mediated phosphorylation of rhodopsin and receptor-mediated activation of GRK2 but not GRK2-mediated phosphorylation of the peptide substrate. Furthermore, the conserved region as well as the C-terminal domain could directly bind rhodopsin in vitro. These results indicate that the C-terminal domain, or more precisely, the conserved region of this domain, is important for enzyme-receptor interaction and that this interaction is required for GRK2 to catalyze receptor phosphorylation.
G蛋白偶联受体激酶（GRKs）的C末端结构域由一个保守区域和一个可变区域组成，并且已表明可变区域可指导胞质酶的膜转运。目前的研究表明，C末端结构域也可能参与主要由保守区域介导的激酶-受体相互作用。GRK2的C末端结构域的截短或该结构域中保守区域的缺失导致其磷酸化视紫红质的能力完全丧失，并且其对受体介导的肽底物磷酸化的敏感性明显降低。相反，GRK2的C末端结构域中βγ亚基结合区域的缺失并未显著改变该酶磷酸化视紫红质的能力。此外，代表GRK2的C末端结构域和保守区域的重组蛋白可以抑制GRK2介导的视紫红质磷酸化以及受体介导的GRK2激活，但不能抑制GRK2介导的肽底物磷酸化。此外，保守区域以及C末端结构域在体外可以直接结合视紫红质。这些结果表明，C末端结构域，或者更确切地说，该结构域的保守区域，对于酶-受体相互作用很重要，并且这种相互作用是GRK2催化受体磷酸化所必需的。
