Chlorophyll Formation in Greening Bean Leaves during the Early Stages

In the evolution of the absorption spectrum of etiolated bean leaves (Phaseolus vulgaris L.) following illumination, a rapid photoconversion of 50% or more of the active protochlorophyllide at room temperature is followed by a shift of the chlorophyll(ide) absorption maximum: C(678)--> -->C(684)-->C(672 nm). Kinetic studies at 2 C and the absence of an isosbestic point provide evidence for an intermediate between C(678) and C(684). A dramatically different evolution is observed following the photoconversion of only 5 to 30% of the active protochlorophyllide at room temperature. C(672) appears within 30 seconds, and no subsequent dark shift occurs during the following 90 minutes. At 0 C, conversion of 5% of the active protochlorophyllide produces a new species, C(676), which converts progressively to C(672) within 10 minutes. We interpret the results in terms of two photochemical steps operating in series for the complete conversion of active protochlorophyllide. Furthermore, there appears to be competition between an irreversible, terminal dark shift and the second light reaction. We propose a scheme based on dimers of protochlorophyllide reduced stepwise to dimers of chlorophyllide in two successive light reactions. The intermediate mixed protochlorophyllide-chlorophyllide dimer absorbs at 676 nm and displays a much faster dissociation to monomers than does the chlorophyllide-chlorophyllide dimer.