Photoionization of ferrocytochrome c by 248 nm laser light and the observation of the early stages of ferricytochrome c unfolding in the nanosecond-to-millisecond timescale.
Photolysis of ferrocytochrome c by 248 nm laser light in aqueous solution at pH 7 generates hydrated electrons (eaq-) by a monophotonic process with quantum yield phi = 0.034. Approximately three-quarters of the eaq- originate from the heme, which is converted from the ferrous to the ferric state in < 100 ns. The conformational changes associated with the change in the redox state of cytochrome c are either not detectable spectrophotometrically or complete in < 100 ns. Also, under conditions where ferrocytochrome c is stable but ferricytochrome c is unfolded (3 M guanidine, pH 7, 40 degrees C), photoionization of ferrocytochrome c generated ferricytochrome c with similar quantum yield. Under these conditions, the lifetime of native ferricytochrome c is 67 microseconds; it decays via two intermediates with lambda max > 410 nm, neither of which is the thermodynamically favored, unfolded form. These species are putatively identified as unfolding intermediates with nonnative iron ligands, similar to those found during folding of ferrocytochrome c. The results suggest that unfolding, like folding, proceeds by intrachain diffusion and ligand exchange.
Candeias
L P
LP
Max-Planck-Institut für Strahlenchemie, Mülheim, Germany. candeias@dsa.mpi-muelheim.mpg.de
Steenken
S
S
D000818:Animals; D003574:Cytochrome c Group / Q000737:chemistry* / Q000528:radiation effects*; D066298:In Vitro Techniques; D007834:Lasers; D010084:Oxidation-Reduction; D010782:Photolysis; D011489:Protein Denaturation / Q000528:radiation effects; D017510:Protein Folding
D003574:Cytochrome c Group