Effects of detergent environments on the photocycle of purified monomeric bacteriorhodopsin.
Time-resolved difference spectra have been obtained for the photocycle of delipidated bacteriorhodopsin monomers (d-BR) in six different detergent micelle environments that were prepared by two new detergent-exchange techniques. A global kinetic analysis of the photocycle spectra for d-BR in each detergent environment was performed. Comparison of these results with those obtained for the photocycle of bacteriorhodopsin in purple membrane (PM) shows that there is one fewer kinetically distinguishable process for monomeric BR between the decay of the K intermediate and the rise of the M intermediate. Assuming a sequential pathway occurs in the photocycle, it appears that the equilibrium between the L and M intermediates is reached much more rapidly in the detergent micelles. This is attributed to a more direct interaction between Asp-85 and the proton on the nitrogen of the Schiff base of retinal for BR in the detergents. Equilibrium concentrations of late photocycle intermediates are also altered in detergents. The later steps of the photocycle, including the decay of the M intermediate, are slowed in detergents with rings in their hydrocarbon region. This is attributed to effects on conformational changes occurring during the decay of M and/or other later photocycle intermediates. The lifetime of dark adaptation of light-adapted d-BR in different detergent environments increases in environments where the lifetime of the M intermediate increases. These results suggest that the high percentage of either unsaturated or methyl-branched lipids in PM and the membranes of other retinal proteins may be important for their effective functioning.
Milder
S J
SJ
Department of Chemistry, University of California, Santa Cruz 95064.
Thorgeirsson
T E
TE
Miercke
L J
LJ
Stroud
R M
RM
Kliger
D S
DS
D001436:Bacteriorhodopsins / Q000302:isolation & purification / Q000378:metabolism* / Q000528:radiation effects; D003902:Detergents / Q000494:pharmacology*; D006217:Halobacterium / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008823:Micelles; D010782:Photolysis; D013053:Spectrophotometry
D003902:Detergents; D008823:Micelles; D001436:Bacteriorhodopsins