The internal lateral pressure of a bilayer has been estimated by numerous investigators. Most of these measurements were made by using the monolayer technique. In our approach, the disk membrane lateral pressure was estimated by assuming that this value is equal to the surface pressure necessary to maintain the transmembrane orientation of rhodopsin. The orientation of rhodopsin at the nitrogen-water interface was determined by using ellipsometry, which can measure the thickness of the film. By examining surface pressure and ellipsometric isotherms of intact and partially hydrolyzed rhodopsin, we have determined that a lateral pressure of 38 mN/m is necessary to give rhodopsin its natural transmembrane orientation and that surface pressures exceeding 45 mN/m lead to the formation of multilayers in the disk membrane film. At 38 mN/m, pure rhodopsin is found to have a molecular area of 2300 Å2.
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