Abstract
A rapid light-induced decrease in cGMP is thought to play a role in regulating the permeability or light sensitivity of photoreceptor membranes. Photo-excited rhodopsin activates a guanine nucleotide-binding protein (G-protein) by catalyzing the exchange of bound GDP for GTP. This G-protein X GTP complex activates the phosphodiesterase resulting in a decrease in cGMP concentration. We have observed two processes in vitro which may be relevant for the regulation of G-protein activation. First, we have found that free GDP binds to G-protein with an affinity similar to that of GTP. These two nucleotides appear to compete for a common site. Since G-protein X GDP does not activate phosphodiesterase, light-induced changes in the GTP/GDP ratio known to occur on illumination may serve to reduce G-protein activation and hence reduce phosphodiesterase activation. Second, addition of cGMP in the presence of equimolar GTP and GDP causes GTP binding to G-protein to be enhanced compared to GDP binding. This effect increases as the cGMP concentration is increased from 0.05 to 2 mM. Thus, light-induced decreases in cGMP concentration may also act as a feedback control in reducing G-protein activation. One or both of these processes may be involved in the desensitization (light adaptation) of rod photoreceptors.
Highlights
The G-protein found in vertebrate rods, which is functionally and structurally analogous to G-proteins found in the
The enzymes that regulate a light-induced drop in cGMP hormone-sensitive adenylate cyclase system (Manningand levels in vertebrate rod photoreceptor cells are thesubject of Gilman, 1983),is comprised of three subunits with molecular increasing interest because of accumulating evidence that masses of39 kDa (a),36 kDa (p), 10 kDa (7)(Kuhn, 1980; cGMP plays an important role in visual transduction Baehr et al, 1982; Fung, 1983)
The open (GDP) and closed function of the reciprocal of the nucleotide concentration.Eachpoint (GTP) diamonds indicate light-activated binding in rod outer seg- represents a single determination and the open and closed symbols ments that were preincubated with a monoclonal antibody “4a” to represent two separate experiments.The apparent K, for GTP and the N subunit of the G-protein according to the methods of Hamm GDP are 7 and 12 pm, respectively
Summary
Light-induced GTP Binding Is Antagonized by GDP-The top two curves of Fig. 1 demonstrate that G-protein binds both GTP and GDP upon flash illumination bleaching 3 x lo5 rhodopsin molecules/outer segment. The fact that addition of equimolar nonradioactive nucleotide causes approximately a 50% inhibition of binding of the other (radioactive) nucleotide suggests that under these conditions the light-activated G-protein does not prefer GTP over GDP.
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