Heterogeneity in regeneration of bacteriorhodopsin from bacterio-opsin and all-trans retinal at high temperatures: implications for dynamic structural fluctuations.

Measurements of regeneration kinetics were performed in order to investigate the regeneration mechanisms of bacteriorhodopsin (bR) from thermally unfolded bacterio-opsin (bO) and all-trans retinal. Regeneration kinetics data were successfully fitted to a single exponential function when regeneration was performed at 25 degrees C after incubation at high temperatures. Conversely, the process of regeneration after the addition of retinal to bO at high temperatures occurred at two different rate constants.

Regeneration of bacteriorhodopsin from thermally unfolded bacterio-opsin and all-trans retinal at high temperatures.

The temperature dependence of regeneration of bacteriorhodopsin (bR) from its apoprotein, bacterio-opsin (bO), and all-trans retinal was investigated using two different procedures to probe the structural properties of bO at high temperatures. Regeneration experiments performed at 25 degrees C after incubation of bO within the temperature range of 35-75 degrees C indicate that irreversible thermal unfolding begins at 50 degrees C. When bO is incubated for one hour and mixed with retinal at the same elevated temperatures, however, a greater extent of regeneration to bR occurs, even at temperatures ranging from 50 to 65 degrees C.