Size and shape of bovine interphotoreceptor retinoid-binding protein by electron microscopy and hydrodynamic analysis.
Individual molecules of interphotoreceptor retinoid-binding protein (IRBP), a protein likely to be important in the visual cycle, were visualized by means of electron microscopy. IRBP was coated with a very thin layer of tungsten and photographed by dark-field imaging. IRBP is seen to be a flexible, elongated molecule about 24 nm in length by 3-4 nm in width (statistical modes). These dimensions agree very well with those calculated from the frictional ratio obtained from sedimentation data. Approximately half of these rod-shaped IRBP molecules are straight, and half are bent in the middle, usually with an angle of 60-90 degrees between the two arms. A representation of IRBP as a bendable string of beads yields calculations of dimensions and of hydrodynamic parameters consistent with the electron microscopic and sedimentation data; the sedimentation coefficients derived from this representation are nearly insensitive to molecular bending. When IRBP is bound to saturating amounts of its endogenous ligands, all-trans- or 11-cis-retinol, its sedimentation behavior is unchanged, and the same types of particles are visualized by electron microscopy as with the free protein; however, a greater proportion of the molecules are bent. Deglycosylation of IRBP (with peptide:N-glycosidase F) results in a somewhat smaller molecule that retains its rod-like shape, as shown by gel filtration and sedimentation data. The results indicate that IRBP is an elongated molecule and suggest that a structural change may occur upon ligand binding.
Adler
A J
AJ
Eye Research Institute of Retina Foundation, Boston, Massachusetts 02114.
Stafford
W F
WF
Slayter
H S
HS
D000818:Animals; D002417:Cattle; D002850:Chromatography, Gel; D005136:Eye Proteins; D008854:Microscopy, Electron; D010786:Photoreceptor Cells / Q000378:metabolism*; D011487:Protein Conformation; D012177:Retinol-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism; D014801:Vitamin A / Q000378:metabolism; D000099214:Retinol-Binding Proteins, Interstitial
D005136:Eye Proteins; D012177:Retinol-Binding Proteins; D014801:Vitamin A; D000099214:Retinol-Binding Proteins, Interstitial