Light and GTP dependence of transducin solubility in retinal rods (original) (raw)
Abstract
The physical origin and functional significance of the near infra-red light scattering changes observable upon flash illumination of diluted suspensions of magnetically oriented, permeabilised frog retinal rods has been reinvestigated with particular attention paid to the degree with which transducin remains attached to the membrane. In the absence of GTP, the so called “binding” signal is shown to include two components of distinctive origins, widely different kinetics, and whose relative amplitudes depend on the dilution of the suspension and resulting detachment of transducin from the disc membrane. The fast component is a consequence of the fast interaction between photoexcited rhodopsin (R*) and the transducin remaining on the membrane. Its kinetics monitors a structural modification of the discs caused by a change in electrostatic interaction between closely packed membranes upon the formation of R*-T complexes. The slow component monitors the slow rebinding to the membrane and possible subsequent interaction with excess R* of _T_-GDP which, in spite of its low solubility, had eluted into solution given the high dilution of the permeated rods. In the presence of GTP, the so called “dissociation” signal includes a fast, anisotropic “release” component that specifically monitors the release into the interdiscal space of T α-GTP formed from the membrane-bound pool, and a slower isotropic “loss” component monitoring the leakage from the permeated rod of the excess T α-GTP which did not interact with the cGMP phosphodiesterase. The amplitudes of both components depend exclusively on the membrane bound T-GDP pool. The kinetics of the “loss” component is limited by the size and degree of permeation of the rod fragments, rather than by the dissociation rate of T α-GTP from the membrane.
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Abbreviations
ROS:
rod outer segment
R:
rhodopsin
R*:
photoactivated rhodopsin
T, _T_-GDP, T α-GDP, T α-GTP, T βγ:
transducin and its various forms
T mb, T sol: T αβγ:
bound to membrane or soluble
PDE:
cGMP-phosphodiesterase
GTP:
guanosine 5′-triphosphate
GDP:
guanosine 5′-diphosphate
GDP βS:
guanosine 5′-O-(2-thiodiphosphate)
cGMP:
guanosine-3′-5′ cyclic-monophosphate
DTT:
dithiothreitol
HEPES:
4-(2-hydroxyethyl)-1-piperazine-ethane sulfonic acid
TRIS:
Tris (hydroxymethyl)aminomethane
SDS:
sodium dodecyl sulfate
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Authors and Affiliations
- Laboratoire de Biophysique Moléculaire et Cellulaire (Unité Associée 520 du CNRS), Fédération de Biologie DRF, CENG, BP 85, F-38041, Grenoble, France
F. Bruckert, T. Minh Vuong & M. Chabre
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- F. Bruckert
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Bruckert, F., Minh Vuong, T. & Chabre, M. Light and GTP dependence of transducin solubility in retinal rods.Eur Biophys J 16, 207–218 (1988). https://doi.org/10.1007/BF00261263
- Received: 11 April 1988
- Accepted: 07 July 1988
- Issue Date: October 1988
- DOI: https://doi.org/10.1007/BF00261263