Interconversion of metarhodopsins I and II: a branched photointermediate decay model.
Flash photolysis experiments designed to monitor the establishment of the metarhodopsin I to metarhodopsin II equilibrium are interpreted according to a branched model in which two spectrally indistinguishable but kinetically distinguishable forms of metarhodopsin II are postulated to exist in equilibrium with a common pool of metarhodopsin I. This interpretation arises from the consistent requirement for at least three exponentials for a valid description of the observed growth of absorbance at 380 nm following bleaching of bovine rhodopsin in rod outer segment disk membranes. Analysis of the 380-nm transient absorbance data permitted direct determination of the five physically interpretable individual rate constants of the model. This analysis represents a more explicit interpretation of kinetic data than that employed in earlier experiments of this kind, which involved estimating only apparent rates and apparent amplitudes of discrete multiexponential functions. The 380-nm absorbance contributions of all relevant species contributing to the observed dynamic absorbance change were accounted for simultaneously during nonlinear least-squares estimation of the model rate parameters. Analysis of deconvoluted equilibrium spectra acquired from samples identical with those used in the kinetics experiments confirmed the metarhodopsin I-metarhodopsin II equilibrium constants, Keq, derived from the dynamic analyses. It is shown that Keq varies from 1.28 at 10 degrees C to 7.3 at 37 degrees C and that approximately 90% of the metarhodopsin II present is in the form of metarhodopsin IIslow over the temperature range 10-37 degrees C. A physical interpretation of this decay model is discussed in the context of a distribution of metarhodopsin II structural and energetic states.
Straume
M
M
Department of Biochemistry, University of Virginia Health Sciences Center, Charlottesville 22908.
Mitchell
D C
DC
Miller
J L
JL
Litman
B J
BJ
D002462:Cell Membrane / Q000187:drug effects; D002784:Cholesterol / Q000494:pharmacology*; D007700:Kinetics; D008051:Lipid Bilayers / Q000378:metabolism*; D008956:Models, Chemical; D010713:Phosphatidylcholines; D010782:Photolysis; D012243:Rhodopsin / Q000031:analogs & derivatives* / Q000378:metabolism; D013053:Spectrophotometry; D013329:Structure-Activity Relationship; D013696:Temperature; D013816:Thermodynamics
D008051:Lipid Bilayers; D010713:Phosphatidylcholines; C013110:metarhodopsins; D012243:Rhodopsin; D002784:Cholesterol