Amino acid residues in GRK1/GRK7 responsible for interaction with S-modulin/recoverin.
GRK1 is a visual pigment kinase in rods and is essential for inactivation of light-activated rhodopsin. The GRK1 activity is inhibited by binding of the Ca(2+)-bound form of S-modulin/recoverin. We previously identified the S-modulin/recoverin site to interact with GRK1. In the present study, we identified its counterpart in GRK1. We synthesized 29 of GRK1 or GRK7 partial peptides that cover the entire sequence of GRK1/GRK7, and examined whether these peptides inhibit S-modulin/recoverin activity most probably by preoccupying the binding site for GRK1. The inhibition was the greatest with the N-terminal peptide (p1, aa 3-23 in GRK7). On mutation of each of eight amino acid residues highly conserved in the p1 region of more than 10 orthologs, the inhibition was significantly reduced in the mutation of Leu(6), Asn(12) and Tyr(15). We further examined the binding of the peptides, including mutated ones, to S-modulin/recoverin with a resonance mirror biosensor. The binding correlated well with the degree of the inhibition by a peptide. The inhibition, therefore, seemed to be due to a direct binding of the kinase peptide to the binding site of active S-modulin/recoverin. A GRK1 region close to its C-terminus also seemed to be the binding site for S-modulin/recoverin.
Torisawa
Aya
A
Graduate School of Frontier Biosciences, Osaka University, Osaka, Japan.
Arinobu
Daisuke
D
Tachibanaki
Shuji
S
Kawamura
Satoru
S
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D051606:G-Protein-Coupled Receptor Kinase 1 / Q000737:chemistry* / Q000378:metabolism*; D054768:G-Protein-Coupled Receptor Kinases / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D051603:Recoverin / Q000737:chemistry / Q000378:metabolism*
D000596:Amino Acids; D051603:Recoverin; D051606:G-Protein-Coupled Receptor Kinase 1; C496764:GRK1 protein, human; C111571:GRK7 protein, human; D054768:G-Protein-Coupled Receptor Kinases; D002118:Calcium