Characterization of a truncated form of arrestin isolated from bovine rod outer segments.
The inactivation of photolyzed rhodopsin requires phosphorylation of the receptor and binding of a 48-kDa regulatory protein, arrestin. By binding to phosphorylated photolyzed rhodopsin, arrestin inhibits G protein (Gt) activation and blocks premature dephosphorylation, thereby preventing the reentry of photolyzed rhodopsin into the phototransduction pathway. In this study, we isolated a 44-kDa form of arrestin, called p44, from fresh bovine rod outer segments and characterized its structure and function. A partial primary structure of p44 was established by a combination of mass spectrometry and automated Edman degradation of proteolytic peptides. The amino acid sequence was found to be identical with arrestin, except that the C-terminal 35 residues (positions 370-404) are replaced by a single alanine. p44 appeared to be generated by alternative mRNA splicing, because intron 15 interrupts within the nucleotide codon for 369Ser in the arrestin gene. Functionally, p44 binds avidly to photolyzed or phosphorylated and photolyzed rhodopsin. As a consequence of its relatively high affinity for bleached rhodopsin, p44 blocks Gt activation. The binding characteristics of p44 set it apart from tryptic forms of arrestin (truncated at the N- and C-termini), which require phosphorylation of rhodopsin for tight binding. We propose that p44 is a novel splice variant of arrestin that could be involved in the regulation of Gt activation.
Palczewski
K
K
Department of Ophthalmology, University of Washington, Seattle 98195.
Buczylko
J
J
Ohguro
H
H
Annan
R S
RS
Carr
S A
SA
Crabb
J W
JW
Kaplan
M W
MW
Johnson
R S
RS
Walsh
K A
KA
D000595:Amino Acid Sequence; D000818:Animals; D000941:Antigens / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D019393:Arrestin; D002417:Cattle; D005136:Eye Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D015151:Immunoblotting; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation; D010782:Photolysis; D012243:Rhodopsin / Q000378:metabolism; D012374:Rod Cell Outer Segment / Q000737:chemistry*; D014357:Trypsin / Q000378:metabolism
D000941:Antigens; D019393:Arrestin; D005136:Eye Proteins; D010446:Peptide Fragments; D012243:Rhodopsin; D014357:Trypsin