Biosynthesis of chlorophyll from protochlorophyll(ide) in green plant leaves.

Using spectral methods, the biosynthesis of protochlorophyll(ide) and chlorophyll(ide) in green plant leaves was studied. The main chlorophyll precursors in the green leaves (as in etiolated leaves) were photoactive photocholorophyll(ide) forms Pchl(ide)655/650(448) and Pchl(ide)653/648(440). The contributions into Chl biosynthesis of the shorter-wavelength precursor forms, which were accumulated in darkened green leaves as well, were completely absent (of Pchl(ide) 633/628(440)) or insignificant (of Pchl(ide) 642/635(444)).

Biosynthesis of chlorophyll of photosystem II reaction centers in plant leaves in early stages of etiolation.

Spectral methods were used to study the sequences of chlorophyll biosynthesis reactions in etiolated pea, bean, and maize plants in early stages (3-4 days) of growth. For these juvenile plants, along with the reaction chain known for mature (7-9 day-old) plants, a new reaction chain was found which started with phototransformation of the long-wavelength form PChld 686/676 into PChld 653/648. (PChld 653/648 differs from the main known precursor form PChld 655/650). The subsequent photoreduction of PChld 653/648 leads to the formation of Chld 684/676, which is transformed into Chl 688/680 in the course of a dark reaction. After completion of this reaction, fast (20-30 sec) quenching of the fluorescence of the reaction product is observed with the formation of non-fluorescent Chl 680. The reaction accompanied by pigment fluorescence quenching is absent in pea mutants with depressed function of Photosystem II reaction centers. This suggests that the newly found reaction chain leads to the formation of chlorophyll of the Photosystem II reaction center.

The final stage of chlorophyll biosynthesis and formation of reaction centres of photosystems.

The survey reviews Russian studies of the final stage of chlorophyll biosynthesis and the role of A. A. Krasnovsky (Sr.) in the development of this direction. The current state of the problem is considered. The results of studies carried out by Krasnovsky's followers at Moscow State University are summarized. Schemes of the pathways of chlorophyll and pheophytin biosynthesis, biogenesis of pigment complexes of the two photosystems of photosynthesis proposed by those scientists are presented.

Photoconversion of long-wavelength protochlorophyll native form Pchl 682/672 into chlorophyll Chl 715/696 in Chlorella vulgaris B-15.

By spectral methods, the final stages of chlorophyll formation from protochlorophyll (ide) were studied in heterotrophic cells of Chlorella vulgaris B-15 mutant, where chlorophyll dark biosynthesis is inhibited. It was shown that during the dark cultivation, in the mutant cells, in addition to the well-known protochlorophyll (ide) forms Pchlide 655/650, Pchl(ide) 640/635, Pchl(ide) 633/627, a long-wavelength protochlorophyll form is accumulated with fluorescence maximum at 682 nm and absorption maximum at 672 nm (Pchl 682/672). According to the spectra measured in vivo and in vitro, illumination of dark grown cells leads to the photoconversion of Pchl 682/672 into the stable long wavelength chlorophyll native form Chl 715/696. This reaction was accompanied by well-known photoreactions of shorter-wavelength Pchl (ide) forms: Pchlide 655/650→Chlide 695/684 and Pchl (ide) 640/635→Chl (ide) 680/670. These three photoreactions were observed at room temperature as well as at low temperature (203-233 K).

Light-regulated pigment interconversion in pheophytin/chlorophyll-containing complexes formed during plant leaves greening.

Upon illumination of etiolated maize leaves the photoconversion of protochlorophyllide Pchlide 655/650 into chlorophyllide Chlide 684/676 was observed. It was shown that chlorophyllide Chlide 684/676 in the dark is transformed into pheophytin Pheo 679/675 and chlorophyll Chl 671/668 by means of two parallel reactions, occurring at room temperature: Chlide 684/676. The formed pheophytin Pheo 679/675 was unstable and in the dark was transformed into chlorophyll Chl 671/668 in a few seconds: Pheo 679/675 → Chl 671/668. The last reaction is reversed by the light: Chl/668 Pheo 679/675. Thus, on the whole in the greening etiolated leaves this process occurs according to the following scheme:The observed light-regulated interconversion of Mg-containing and Mg-free chlorophyll analogs is activated by ATP and inhibited by AMP.

Photoinduced formation of pheophytin/chlorophyll-containing complexes during the greening of plant leaves.

Illumination of etiolated maize leaves with low-intensity light produces a chlorophyll/pheophytin-containing complex. The complex contains two native chlorophyll forms Chl 671/668 and Chl 675/668 as well as pheophytin Pheo 679/675 (with chlorophyll/pheophytin ratio of 2/1). The complex is formed in the course of two successive reactions: reaction of protochlorophyllide Pchlde 655/650 photoreduction resulted in chlorophyllide Chlde 684/676 formation, and the subsequent dark reaction of Chlde 684/676 involving Mg substitution by H2 in pigment chromophore and pigment esterification by phytol. Out data show that the reaction leading to chlorophyll/pheophytin-containing complex formation is not destructive. The reaction is in fact biosynthetic, and is competitive with the known reactions of biosynthesis of the bulk of chlorophyll molecules. The relationship between chlorophyll and pheophytin biosynthesis reactions is controlled by temperature, light intensity and exposure duration.The native complex containing pheophytin a and chlorophyll a is supposed to be a direct precursor of the PS II reaction centre in plant leaves.

Low temperature phototransformations of protochlorophyll(ide) in etiolated leaves.

By methods of difference and derivative spectroscopy it was shown that in etiolated leaves at 77 K three photoreactions of P650 protochlorophyllide take place which differ in their rates and positions of spectral maxima of the intermediates formed in the process: P650→R668, P650→R688, and P650→R697. With an increase of temperature up to 233 K, in the dark, R688 and R697 are transformed into the known chlorophyllide forms C695/684 and C684/676, while R668 disappears with formation of a shorter wavelength form of protochlorophyllide with an absorption maximum at 643-644 nm.Along with these reactions, at 77 K phototransformations of the long-wave protochlorophyllide forms with absorption maxima at 658-711 nm into the main short-wave forms of protochlorophyllide are observed. At 233 K in the dark this reaction is partially reversible. This process may be interpreted as a reversible photodisaggregation of the pigment in vivo.The mechanism of P650 reactions and their role in the process of chlorophyll photobiosynthesis are discussed.

The role of a long-wavelength pigment form in the chlorophyll biosynthesis.

Photoconversion of protochlorophyllide650 form was observed in etiolated leaves illuminated with long-wavelength-690 nm-light. This process showed Shibata shift and was found to have a strong temperature dependence between 20 and -40°C. The low rate of reaction, the strong temperature dependence and calculations on the spectral overlap integral of absorption and fluorescence bands in this spectral region indicate that the phototransformation of the 650 nm form of protochlorophyllide may be caused by a back energy migration from a long-wavelength pigment form absorbing around 690 nm; this pigment form is probably a long-wavelength form of protochlorophyll/ide.

[Energy migration in a pigmented protochlorophyllide complex]. / Migratsiia énergii v pigmentnom komplekse protokhlorofillida.

Data are analysed concerning energy migration between protochlorophyllide and chlorophyllide formed in etiolated leaves. It is found that phototransformation of the precursor proceeds in the pigment complexes similar to the multicentric photosynthetic unit. The complexes comprise no more than 26 molecules of the active form of protochlorophyllide P650 and two molecules of minor forms of P628 and P639 on the average with mean intermolecular distances 3 nm.

[Energy migration in photoactive complexes of chlorophyll precursor in etiolated leaves and spectroscopic characteristics of pigment forms]. / Migratsiia énergii v fotoaktivnykh kompleksakh predshestvennika khlorofilla v etiolirovannykh list'iakh i spektroskopicheskie kharakteristiki pigmentnykh form

Migration energy between three forms of the precursor of chlorophyll and chlorophyllide are measured and calculated. Absorption coefficients, fluorescence yields of pigment forms and dimensions of native complexes (20-22 molecules) are calculated.