Light-dependent binding of G-protein to outer segment membranes of toad photoreceptors.
Light-dependent binding of G-protein to outer segment membranes of toad photoreceptors.
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DOI：
10.1085/jgp.88.5.675
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发表时间：
1986-11
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影响因子：
--
通讯作者：
Baehr W
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文献类型：
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作者：
Mangini NJ;Pepperberg DR;Baehr W
本文分析了蟾蜍（Bufo marinus）视网膜光感受器外节盘膜中g蛋白结合的光依赖性变化。分离的完整视网膜，在23 +/- 1℃的含氧林格氏溶液中培养，在不同的光照条件下，然后在黑暗中培养特定的时间。然后将视网膜冷冻（0-4℃），分离受体外段（ROS）。通过对低渗培养基中不含GTP和含GTP的低渗培养基（分别为H和HG提取物）中提取的G α和G β与ROS膜的结合进行定量分析。对于光照后立即冷藏进行分析的视网膜，G结合的程度（HG提取物中G α和β的相对丰度）随着视觉色素的漂白程度而增加。在漂白大于或等于30%后，观察到接近最大的结合。在漂白约70%后，随着在黑暗中孵育时间的增加，结合程度逐渐下降到未漂白视网膜的低水平。半完成下降所需的时间约为10(3)s。随着强光照射时间的增加，G结合也从一个快速发展的峰值逐渐下降。从之前的电生理数据来看，我们的研究结果表明，视杆细胞的持续漂白脱敏并不依赖于g蛋白被漂白的视觉色素“紧密结合”（固定）的持续状态。
Light-dependent changes in the binding of G-protein were analyzed in outer segment disk membranes obtained from photoreceptors of the toad (Bufo marinus) retina. Isolated, intact retinas, incubated in oxygenated Ringer's solution at 23 +/- 1 degree C, were subjected to various conditions of illumination and then incubated in darkness for specified periods. The retinas were then chilled (0-4 degrees C) and the receptor outer segments (ROS) were isolated. Binding of the alpha- and beta-subunits of G-protein to the ROS membranes was analyzed by quantitating G alpha and G beta extracted from the membranes with hypotonic medium lacking GTP vs. hypotonic medium containing GTP (H and HG extracts, respectively). For retinas illuminated and then immediately chilled for analysis, the extent of G binding (relative abundance of G alpha, beta in the HG extract) increased with the extent of bleaching of the visual pigment. Near-maximal binding was observed after bleaches of greater than or equal to 30%. With an increasing period of incubation in darkness after approximately 70% bleaching, the extent of binding declined gradually to low levels characteristic of unbleached retinas. The period required for half-completion of the decline was approximately 10(3) s. A gradual decline in G binding, from a rapidly developing peak value, was also observed with an increasing period of exposure to intense light. Viewed in the context of previous electrophysiological data, our results indicate that sustained bleaching desensitization of the rods does not depend upon a persisting state of "tight binding" (immobilization) of G-protein by bleached visual pigment.