Assessment of physiological integrity of sonicated retinal rod membranes

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Abstract
As the size of rod membrane fragments was reduced by sonication or by addition of the detergent Emulphogene, the intensity of the circular dichroism (CD) bands (210 and 221 nm) increased progressively with a blue shift in position. The intensity of the visible CD bands (340 and 495 nm) was also increased by sonication. Since the intensity increase of the CD bands was related to a reduction in turbidity, the anomalous CD features of intact membranes could be attributed to optical artifacts caused by the particulate nature of the material.Because the magnitude of the CD bands at 221 nm and 340 nm was essentially identical for the sonic suspension and detergent‐clarified solution, the adequacy of sonic suspensions can be assured by checking whether detergent affects the intensity of these bands.Suspensions of sonicated rod membranes, purified on Agarose, contained vesicles of 112 nm in average diameter. The morphology and size of the vesicles did not change upon photobleaching of rhodopsin. The vesicles retained such rod membrane properties as conformational insensitivity to photobleaching of the retinal chromophore, thermal stability, and pigment regenerability. Thus, the physiological integrity of rod membranes was maintained by the sonicated vesicles.From the most reliable estimate of the molecular ellipticity at 221 nm, the helical content of membrane‐bound rhodopsin was determined to be approximately 47%.