Abstract
A new protocol for the preparation of recombinant phytochromes results in significantly higher yields which, for the first time, have made kinetic studies possible. Flash photolysis with nanosecond laser excitation reveals that, in recombinant and native phytochromes, the decay kinetics of the primary photoproducts I 700 i and the kinetics of the formation of the P fr form are similar. Phycocyanobilin-containing recombinant phytochrome, however, shows only a monoexponential decay of the I 700 intermediate with a time constant of approximately 90 μs, and a biexponential formation of the P fr form, albeit with time constants (approximately 13 and 100 ms) somewhat shorter than those from native phytochrome. Thus the seemingly small structural modification of the chromophore (substitution of the native vinyl for an ethyl group) has a profound influence on the availability of protein conformational rearrangement pathways. The result is therefore of general interest in chromoprotein dynamics.
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