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H R Saibil

Publications and source records attributed to H R Saibil.

43 records · Page 3Linked to original sources
PubMed1991

Effects of calcium on light-activated GTP-binding proteins in squid photoreceptor membranes.

1. At least two distinct G-proteins are activated by light in squid photoreceptor membranes, a 45,000 mol. wt cholera toxin substrate and a 40,000 mol. wt pertussis toxin substrate. 2. The light-stimulated GTPase activity is partially inhibited by pretreatment with either toxin and abolished by treatment with both toxins. 3. At 24 degrees C, a free calcium ion concentration of 1 microM inhibits GTPase activity of both toxin substrates and ADP ribosylation by pertussis toxin. 4. This calcium sensitivity of squid G-proteins may be important in interpretation of experimental results on the phosphoinositide or other signalling pathways in squid visual transduction.

Adenosine Diphosphate Ribose↗
PubMed1988

Light- and GTP-activated hydrolysis of phosphatidylinositol bisphosphate in squid photoreceptor membranes.

Light stimulates the hydrolysis of exogenous, [3H]inositol-labeled phosphatidylinositol bisphosphate (PtdInsP2) added to squid photoreceptor membranes, releasing inositol trisphosphate (InsP3). At free calcium levels of 0.05 microM or greater, hydrolysis of the labeled lipid is stimulated up to 4-fold by GTP and light together, but not separately. This activity is the biochemical counterpart of observations on intact retina showing that a rhodopsin-activated GTP-binding protein is involved in visual transduction in invertebrates, and that InsP3 release is correlated with visual excitation and adaptation. Using an in vitro assay, we investigated the calcium and GTP dependence of the phospholipase activity. At calcium concentrations between 0.1 and 0.5 microM, some hydrolysis occurs independently of GTP and light, with a light- and GTP-activated component superimposed. At 1 microM calcium there is no background activity, and hydrolysis absolutely requires both GTP and light. Ion exchange chromatography on Dowex 1 (formate form) of the water-soluble products released at 1 microM calcium reveals that the product is almost entirely InsP3. Invertebrate rhodopsin is homologous in sequence and function to vertebrate visual pigment, which modulates the concentration of cyclic GMP through the mediation of the GTP-binding protein transducin. While there is some evidence that light also modulates PtdInsP2 content in vertebrate photoreceptors, the case for its involvement in phototransduction is stronger for the invertebrate systems. The results reported here support the scheme of rhodopsin----GTP-binding protein----phospholipase C activation in invertebrate photoreceptors.

Animals↗
PubMed1984

A light-stimulated increase of cyclic GMP in squid photoreceptors.

Photoreceptor outer segments isolated from squid retina are known to contain a light-activated GTP-binding protein. Here it is shown that these photoreceptors contain around 0.01 mol cyclic GMP per mol rhodopsin. Adding GTP in the dark stimulates the production of 0.0003-0.001 mol cyclic GMP/mol rhodopsin per min. GTP and light cause a 2-fold faster increase in cyclic GMP. These results show that either (1) squid rhodopsin activates a guanylate cyclase, or (2) there is a constant guanylate cyclase activity and photoexcited rhodopsin inhibits a cyclic GMP phosphodiesterase.

Animals↗
PubMed1984

Squid rhodopsin and GTP-binding protein crossreact with vertebrate photoreceptor enzymes.

The activation of photoreceptor GTP-binding protein by rhodopsin was studied in squid photoreceptors and in crossreactions between the squid and bovine proteins. Turbidity changes were observed in the far-red after photoexcitation of rhodopsin with brief flashes and were used to probe interactions between photoreceptor membrane suspensions and soluble protein extracts. Our findings are squid photoreceptors contain a GTP-binding protein detectable by light- and GTP-sensitive turbidity changes and by limited sequence homology of a 46-kilodalton polypeptide to the alpha-subunit of bovine GTP-binding protein; the squid membranes activate bovine GTP-binding protein qualitatively in the same way as bovine rhodopsin; the 46-kilodalton component is present in a membrane-bound fraction but is more abundant in a crude, soluble fraction of squid rhabdomes, and this soluble fraction can interact with either squid or bovine rhodopsin-containing membranes; light-activated GTPase activities in all of these preparations are consistent with the light-induced turbidity changes. These results show that rhodopsin activation of GTP-binding protein is highly conserved in vertebrate and cephalopod photoreceptors. Since squid rhodopsin is immobilized in precisely ordered microvilli, this suggests that activation of GTP-binding protein in cephalopod photoreceptors occurs in the absence of rhodopsin diffusion. The rhodopsin immobility may be compensated by higher mobility of the soluble GTP-binding protein.

Animals↗
PubMed1979

Orientation of rhodopsin alpha-helices in in retinal rod outer segment membranes studied by infrared linear dichroism.

Frog retinal rod outer segments, oriented by a magentic field, were shown to contain rhodopsin alpha-helical segments preferentially aligned perpendicular to the plane of the disc membrane, by the technique of infrared linear dichroism. Infrared absorption parallel and perpendicular to the rod axes by peptide C parallel to O groups, whose absorption band contains alpha-helical and random coil components at slightly different frequencies, showed positive dichroism centered on the alpha-helix frequence. We conclude that the alpha-helical portion of the protein has an average orientation in the transmembrane direction. Furthermore, infrared spectra of rods in 2H2O Ringer's solution exhibit two distinct peptide amino group absorption bands: the unexchanged N-2H band, which is nondichroic. This implies that the oriented part of the protein is in the lipid bilayer, supporting a model for rhodopsin with a hydrophobic core containing partially oriented alpha-helices and hydrophilic ends consisting of unoriented polypeptide.

Animals↗
PubMedDate not supplied

Projection structure of an invertebrate rhodopsin.

Rhodopsin is the G-protein-coupled membrane receptor that initiates the visual transduction cascade in retinal photoreceptors. In the present study rhodopsin from the dark-adapted retinas of squid (Loligo forbesi) was detergent-extracted, purified, and reconstituted into native squid photoreceptor lipids following proteolytic cleavage of its prolinerich C-terminus. Two-dimensional crystals of C-terminally truncated rhodopsin reconstituted from octyl glucoside solution formed in a p222(1) lattice (a = 44 A, b = 131 A). Electron micrographs of frozen-hydrated crystals were processed and a projection structure to 8 A resolution was calculated. The projection map obtained is very similar to maps previously determined for bovine and frog rhodopsins although the crystal packing of the molecules is quite different. Comparison of the maps shows that the arrangement of alpha-helices in the proteins is very similar despite their great phylogenetic distance; this structure is likely to be present in the whole superfamily of G-protein-coupled receptors. Invertebrate rhodopsins have a large insertion in the helix 5-helix 6 loop. Assignment of an additional density in the squid rhodopsin map to this region supports a previously proposed helix assignment and identifies the end-to-end contacts as helices 1 and 5.

Animals↗