Abstract
This paper deals with the best conditions for preparing rhodopsin particulates by means of sonic treatment of rod outer segments separated from the dark adapted cattle retina. Thorough washing and subsequent sonic treatment of water suspension of rod outer segments gave fairly high optical purities, namely, ca. 0.3 of E400mu/E500mu and ca.3.0 of E280mu/E500mu.As long as rhodopsin articulates were kept at 7°C, the absorption spectra remained fairly stable even 7days after preparation in spite of slightly growing turbidity of solution.
Electron microscopic observation revealed that rhodopsin particulates consisted of globular particles in diameter of 50 Å to 200 Å. When illuminated, the diameter of particulate became larger, probably indicating the tendency of aggregation of particles upon illumination. On the basis of this observation, a model involving the rearrangement of rhodopsin lipo-protein micelle was postulated.
Titration of rhodopsin particulates and the bleached products signified a decreasing tendency in the number of acid titrable groups of the latter which almost reached 15% of total titrable groups at PH 3.01. This result is not consistent with the titration data of rhodopsin digitonin complex. 87 groupswere titrable per mole of rhodopsin in the range of PH 3.75 to 10.0. Also, it seems true that the isoionic point of rhodopsin particulates (6.3-6.5) is shifted remarkably to the alkaline side than that of rhodopsin digitonin complex (5.2-5.3).
This work was mainly performed at Honjo Laboratory, Department of Biology, Faculty of Science, Osaka University. The author is particularly obliged to Professor Ichijiro HONJO for his courtesy in making available the facilities of his laboratory and in preparing this manuscript.
Sincere thanks are also due to Professor Sadayoshi KAMIYA, Dapartment of Ophthalmology, Medical School of Nara, for his generous help in making this accomplishment possible.
The author is also indebted to Dr. Yuzo SEKOGUCHI of Honjo Laboratory for his valuable suggestion and kind help throughout this experiment, and to Dr. Akira TANAKA for his technical contribution in preparing the electron microscopic pictures.
