Subunit composition of the peripherin/rds-rom-1 disk rim complex from rod photoreceptors: hydrodynamic evidence for a tetrameric quaternary structure.
Peripherin/rds and rom-1 are homologous integral membrane protein subunits found as an oligomeric complex at the rim regions of rod and cone photoreceptor outer segment disks. These proteins are essential for the morphogenesis of normal outer segments and have been linked to a variety of human retinal degenerative diseases. Previous studies have suggested that disulfide-linked homodimers of peripherin/rds and rom-1 can associate noncovalently to form higher order structures. We have characterized the hydrodynamic properties of Triton X-100 solubilized peripherin/rds-rom-1 complexes from bovine ROS membranes by gel exclusion chromatography on Sepharose C1-6B and velocity sedimentation through H2O- and D2O-based sucrose gradients. A single hydrodynamic species is observed which has a Stokes radius of 6.2 nm, a sedimentation coefficient (S20,w) of 5.8 S, and a partial specific volume of 0.83 mL/g. From these data the molecular mass of the detergent-peripherin/rds-rom-1 complex is calculated to be 240 kDa. The protein component of this complex is estimated to be 135 kDa, providing direct evidence that the solubilized peripherin/rds-rom-1 complex is a tetramer. The abundance of this complex as measured by competitive ELISA and immunoaffinity purification is approximately 4% of total bovine ROS membrane protein and indicates that peripherin/rds-rom-1 tetramers are present at a relatively high average surface density (ca. 4100/ microns m2) at the rim surfaces of rod outer segment disks.
Goldberg
A F
AF
Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, Canada.
Molday
R S
RS
D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005136:Eye Proteins / Q000737:chemistry* / Q000302:isolation & purification; D007381:Intermediate Filament Proteins / Q000737:chemistry* / Q000302:isolation & purification; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification; D008970:Molecular Weight; D009419:Nerve Tissue Proteins; D064531:Peripherins; D012374:Rod Cell Outer Segment / Q000737:chemistry; D059470:Tetraspanins
D005136:Eye Proteins; D007381:Intermediate Filament Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; C578994:PRPH protein, human; C578997:PRPH2 protein, human; D064531:Peripherins; C513106:ROM1 protein, human; D059470:Tetraspanins