The charge-transfer complex between protochlorophyllide and NADPH: An intermediate in protochlorophyllide photoreduction

A hypothesis describing the mechanism of photoactive protochlorophyllide (P) photoreduction in vivo, relating mainly to the molecular nature of the intermediates, is proposed. The hypothesis is compatible with currently published experimental data. After illumination of etiolated barley leaves at 143 to 153 K, the absorption of P remains essentially unchanged, but a new absorption band at 690 nm is observed. Appearance of this new intermediate enables to distinguish between light and dark stages of the photoconversion reaction. When returned to the higher temperature in the dark, the treated leaves begin accumulating chlorophyllide (Chlide), concomitant with the disappearance of the 690-nm band. The decay time of the excited P (P*) is estimated at 300 ps, which approximates the time constant of photoinduced electron transfer (ET). It is suggested that the charge-transfer complex (CTC) in its ground state (GS) (ground state of CTC formed by the partial (δ) electron transfer), i.e. (p^δ-•••H-D^δ+), between P and NADPH - the electron and proton donor (H-D) - accumulates in the following sequence: P* + H-D → (P*•••H-D)→[(P*•••H-D)↔ (P^-•••H-D⁺)] → ¹(P^-•••H-D⁺)] → ³(P^-•••H-D⁺) → (P^δ-•••H-D^δ+), where an equilibrium state (ES) - [(P*•••H-D)↔(P^-•••H-D ⁺)] - with a lifetime of about 1 to 2 ns, exists between the local excited (LE) and ET states. The existence of a triplet ET state - ³(P^-•••H-D⁺) - is proposed because the time interval between recording of the ES and appearance of the CTC GS (35-250 ns) does not fit the lifetime of the singlet excited complex (exciplex). It is feasible that apart from NADPH, other intermediate proton carriers are contemporaneously involved in the dark reaction (P^δ-•••H-D^δ+) → Chlide, because proton binding to the C₇-C₈ bond in vivo takes place in the trans-configuration. The hydride ion may approach the C₇-C₈ bond from one side by heterolytic fission and an additional proton, donated by the protein group, may be simultaneously added to this bond from the opposite side of the porphyrin nucleus surface.