Development of Photochemical Activity and the Appearance of the High Potential Form of Cytochrome b-559 in Greening Barley Seedlings

Abstract

The development of photochemical activity during the greening of dark-grown barley seedlings (Hordeum vulgare L. cv. Svalofs Bonus) was studied in relation to the formation of the high potential form of cytochrome b-559 (cytochrome b-559HP). Photosynthetie oxygen evolution from leaves was detected at 30 minutes of illumination. The rate of oxygen evolution per gram fresh weight of leaf was as high at 2 to 2.5 hours of greening as at 24 hours or in fully greened leaves. On a chlorophyll basis, the photosynthetic rate at 90 minutes of greening was 80-fold greater than the rate at 45 hours. It is concluded that the majority of photosynthetic units are functional at an early stage of greening, and that chlorophyll synthesis during greening serves to increase the size of the units.Plastids showed substantial photochemical oxygen evolution after a seedling greening time of 1 hour. However, a comparison of the relative activity of leaves and plastids at 2 hours and 24 hours of greening suggests that there was some inactivation of greening plastids during isolation. Appreciable photosystem I activity was observed as early as 15 minutes of greening.The synthesis of cytochrome b-5591p during greening does not correlate with the onset of oxygen evolution. Cytochrome b-559HP was absent from etioplasts and in most preparations of 2-hour plastids. The average amount of cytochrome b-559HP at 2 hours of greening was well below the level needed to provide 1 molecule of the carrier for each functional photosynthetic chain. The results suggest that cytochrome b-559Hp is not essential for oxygen evolution. Cytochrome t, cytochrome be, and the low potential forn of cytochrome b-559 were present in the etioplast. There was little increase in the levels of these cytochromes during 24 hours of greening.In a previous study (5), it was shown that cytochrome f and cytochrome b6, which are components of photosystem I (7), are present in the plastids of dark-grown bean leaves, but cytochrome b-559, a component of photosystem II, is absent. Cytochrome b-559 was detectable after 6 hr of greening. It increased gradually 1 Permanent address: Institute of Genetics, University of Copenhagen, Denmark. on further illumination of the seedlings, but even at 16 hr, the amount of cytochrome b-559 relative to cytcchrome f and cytochrome b6 was well below that of mature chloroplasts. Bendall (2) demonstrated that the absorption band attributed to cytochrome b6 in the earlier work with chloroplasts (7) and etioplasts (5) was due to the presence of two dithionite-reducible cytochromes with a-bands at 563 to 564 nm (cytochrome bQ) and 559 nm (cytochrome b-559Lp), respectively. Plesnicar and Bendall (32) confirmed the observation of Boardman (5) that the high potential form of cytochrome b-559 (cytochrome b-559HP) is absent from etioplasts. In the present work, we have studied the appearance of cytochrome b-559HP in plastids from greening barley seedlings in relation to the onset of oxygen evolution in the leaves and in isolated plastids, and on the ...

A photosystem I mutant in barley (Hordeum vulgare L.)

“…To further localize the observed defect of electron transfer in viridis-n 34, the cytochrome levels in mutant and wild-type chloroplasts were determined from their reduced minus oxidized difference spectra (15). On a chlorophyll basis, the measured levels of cytochromes f, b6, and b559 (Table II) were found to be slightly increased in viridis-n 34 compared with wild-type and the low photosystem I activity observed in viridisn 34 is thus not due to the lack of a cytochrometype electron carrier.…”

“…Quantitative determination of the content of cytochromes f, b559, and b6 in isolated chloroplasts was carried out by reduced minus oxidized difference spectroscopy (15). Ferricyanide was used as the chemical oxidant, sodium dithionite and hydroquinone as chemical reductants.…”

“…Photosynthetic oxygen evolution associated with photosystem II electron transfer was measured by use of an oxygen electrode (Rank Bros., Bottisham, Cambridge, U.K.) (15). Photosystem I was assayed using ascorbate plus TMPD as the donor system and methyl viologen as the acceptor.…”

Isolation and characterization of cytochromeb-559 from chloroplasts and etioplasts of barley

“…Since it remains to be established whether the low and high potential forms of cytochrome b-559 have an identical protein moiety or not, the possibility of the isolation of two different proteins has to be considered. The high potential form of cytochrome b-559 is not present in etioplasts (18,37). Isolation of cytochrome b-559 from this plant material would accordingly be expected to result in the isolation of the protein moiety of an in situ low potential form of cytochrome b-559 Surprisingly, the attempt to isolate cytochrome b-559 from etioplasts using either of the two previous mentioned isolation methods was unsuccessful.…”

“…Nevertheless, studies using mutants of Chlamydomonas indicated that the pool of ascorbate-reducible cytochrome b-559 photooxidizable at 77K is different from the pool photooxidizable in the presence of carbonylcyanide-p-trifluoromethyloxyphenylhydrazone (FCCP) (35). In contrast to chloroplasts, etioplasts contain no cytochrome b-559HP (18,37).…”