Effect of NADPH on formation and decay of human metarhodopsin III at physiological temperatures

Difference absorption spectra were recorded during the formation and decay of metarhodopsin III after sonicated membrane suspensions of rhodopsin were bleached at 37 degrees C. The data were analyzed using SVD, spectral decom position and global exponential fitting. By comparison of the results in th e presence or absence of 70 mu M NADPH and those for bovine or human rhodop sin, a single comprehensive scheme was fit to all the data, including reduc tion of retinal to retinol by the intrinsic retinol dehydrogenase. On the t ime scale studied the mechanism involves two 382 nm absorbing species and t wo 468 nm, absorbing species, supporting the notion that human metarhodopsi n III is not a homogeneous species. The results confirm that metarhodopsin III forms and persists sufficiently long in the human retina under physiolo gical conditions that it could undergo secondary photoisomerization. (C) 20 00 Elsevier Science Ltd. All rights reserved.