Fluorescence lifetime,yield, energy transfer and spectrum in photosynthesis, 1950–1960*

The fluorescence lifetime of chlorophyll agives information about the primary photo-physical events in photosynthesis. Most of the light energy absorbed by chlorophylls is utilized for photochemistry. There are two main additional pathways competing for the absorbed light energy: fluorescence and radiationless internal conversion (heat). Only a few percent of the absorbed energy proceeds along these two pathways. This historical minireview focuses on the first direct measurements of the lifetime of chlorophyll fluorescence, the time it takes to transfer energy from phycoerythrin to chlorophyll a, and the discovery of the fluorescence band at 720 nm (F720; then attributed to a dimer of chlorophyll). These works were carried out during the the late 1950s to the early 1960s in the laboratory of Professor Eugene Rabinowitch at the University of Illinois, Urbana-Champaign [Brody (1995) Photosynth Res 43: 67–74]. This Minireview is dedicated to Professor Eugene Rabinowitch (1901–1973), mentor of the author (Steve Brody) as well as of the editor, and authors classmate (Govindjee). The career and contributions of Eugene Rabinowitch are available in a dedication by Bannister (1972). This revised version was published online in June 2006 with corrections to the Cover Date.     

Changing concepts about the distribution of Photosystems I and II between grana-appressed and stroma-exposed thylakoid membranes

Thylakoid membranes of higher plants and some green algae, which house the light-harvesting and energy transducing functions of the chloroplast, are structurally unique. The concept of the photosynthetic unit of the 1930s (Robert Emerson, William Arnold and Hans Gaffron), needing one reaction center per hundreds of antenna molecules, was modified by the discovery of the Enhancement effect in oxygen evolution in two different wavelengths of light (Robert Emerson and his coworkers) in the late 1950s, followed by the 1960 Z scheme of Robin Hill and Fay Bendall. It was realized that two light reactions and two pigment systems were needed for oxygenic photosynthesis. Changing ideas about the distribution of Photosystem II (PS II) and PS I between the green-appressed and stroma-exposed thylakoid membrane domains, which led to the concept of lateral heterogeneity, are discussed. This revised version was published online in June 2006 with corrections to the Cover Date.     

The discovery and function of plastocyanin: A personal account

A brief autobiographical account is presented of the early research that led to the discovery of the copper protein plastocyanin and the identification of its function as an electron carrier in plant photosynthesis. A discussion follows of different approaches employed for the determination of the functional site of plastocyanin in relation to cytochrome f. A summary is provided of a heated controversy about the involvement of two or three light reactions in photosynthesis and an experiment is described that has contributed to resolution of the controversy through the identification of the functional site of plastocyanin. An early history of photosynthesis research in Japan is also discussed.Abbreviation DCIP 2,6-dichlorophenolindophenol Invited and edited by Govindjee.     

What governs the reaction center excitation wavelength of photosystems I and II?

The sun’s spectrum harvested through photosynthesis is the primary source of energy for life on earth. Plants, green algae, and cyanobacteria—the major primary producers on earth—utilize reaction centers that operate at wavelengths of 680 and 700 nm. Why were these wavelengths “chosen” in evolution? This study analyzes the efficiency of light conversion into chemical energy as a function of hypothetical reaction center absorption wavelengths given the sun’s spectrum and the overpotential cost associated with charge separation. Surprisingly, it is found here that when taking into account the empirical charge separation cost the range 680–720 nm maximizes the conversion efficiency. This suggests the possibility that the wavelengths of photosystem I and II were optimized at some point in their evolution for the maximal utilization of the sun’s spectrum.     

Photochemical Effects of Sunlight

The importance of sunlight in bringing about not only photosynthesis in plants, but also other photochemical effects, is reviewed. More effort should be devoted to photochemical storage of the sun's energy without the living plant. There is no theoretical reason to believe that such reactions are impossible. Ground rules for searching for suitable solar photochemical reactions are given, and a few attempts are described, but nothing successful has yet been found. Future possibilities are suggested. Photogalvanic cells which convert sunlight into electricity deserve further research. Eugene Rabinowitch has been an active pioneer in these fields.     

Equilibrium constants and photosynthetic enhancement

The linear (Myers) and nonlinear (Joliot-Kok) models of photosynthesis were used to predict enhancement. With an apparent equilibrium constant of 1000 between the two light reactions, both models predict a minimal enhancement of about 1.0. However, with an equilibrium constant of 5, both models predict a minimal enhancement significantly greater than 1.0. Consequently, with an equilibrium constant of 5, neither model can account for the observed enhancement values of 1.0 near 685 nanometers in Chlorella. Also, with an equilibrium constant of 5, enhancement significantly greater than 1.0 is predicted between two short wavelengths or between two long wavelengths; neither is observed.     

Action Spectra of Photosynthetic Activities of Chlorella ellipsoidea

• 1) Suspensions of Chlorella show an even stronger light scattering than suspensions of chloroplasts of spinach. The bands of absorption are thus broadened and, at higher concentrations, moved to lower wave-lengths. The intensity of the photosynthesis closely follows the curves of light scattering, a fact partly explaining the high efficiency of green light. Calculated per unit thermoelectrically measured incident energy the action spectrum shows bands at 660–670 nm and c. 500 nm and a comparatively high level of the whole region 500–560 nm.
• 2) Flash experiments show the existence of a steady state carotene/xanthophyll that is moved to reduction (c/x > 1) in blue and green light and to oxidation (c/x < 1) in red light. All experiments point to the existence of two light reactions, the first one involving excitation of carotenoids, with ferredoxin-TPN as acceptor, the second one involving excitation of chlorophyll, with the cytochrome system of the chloroplasts acting as donors of electrons and thus completing an energy converting circulation between pigments and enzyme systems.
• 3) The operation of combined light reactions appears also from experiments with simultaneous or succedaneous illumination with monochromatic light of different wave-lengths. Some effects may be explained from separate excitations of carotenoids and chlorophylls, others may depend on still unknown photic reactions.
• 4) The action spectrum in ultrared shows a positive band at c. 900 nm but no or only very small effects in the region 950–1400 nm. Ultrared radiation has on the other hand an enhancing effect on the light excitation in the visible spectrum. A combination of infrared and visible radiation shows a roughly linear relation between incident energy and photosynthetic effect.
• 5) All experiments were performed in the region of linear relation between intensity of incident light and O₂-production. Induced effects of combined monochromatic regions show a very rapid initial change in the steady states that in one or two minutes simmers down to a balanced state of continued photosynthesis. No change was observed in the total quantity of the pigments.

     

Evidences from Action Spectra for a Specific Participation of Chlorophyll b in Photosynthesis

Rate of oxygen evolution in photosynthesis was measured as the current from a polarized platinum electrode covered by a thin layer of Chlorella. The arrangement gave a reproducibly measurable rate of photosynthesis proportional to light intensity at the low levels used and gave rapid response to changes in illumination. Two phenomena have been explored. The Emerson effect was observed as an enhancement of photosynthesis in long wavelength red light (700 mµ) when shorter wavelengths were added. Two light beams of wavelengths 653 and 700 mµ when presented together gave a photosynthetic rate about 25 per cent higher than the sum of the rates obtained separately. Large and reproducible transients in rate of oxygen evolution were observed accompanying change in illumination between two wavelengths adjusted in intensity to support equal steady rates of photosynthesis. The transients were found not to be specifically related to long wavelength red light. Both enhancement and the transients have identical action spectra which are interpreted as demonstrating a specific photochemical participation of chlorophyll b.     

The controversy over the minimum quantum requirement for oxygen evolution

During the early- to mid-twentieth century, a bitter controversy raged among researchers on photosynthesis regarding the minimum number of light quanta required for the evolution of one molecule of oxygen. From 1923 until his death in 1970, Otto Warburg insisted that this value was about three or four quanta. Beginning in the late 1930s, Robert Emerson and others on the opposing side consistently obtained a value of 8–12 quanta. Warburg changed the protocols of his experiments, sometimes in unexplained ways, yet he almost always arrived at a value of four or less, except eight in carbonate/bicarbonate buffer, which he dismissed as “unphysiological”. This paper is largely an abbreviated form of the detailed story on the minimum quantum requirement of photosynthesis, as told by Nickelsen and Govindjee (The maximum quantum yield controversy: Otto Warburg and the “Midwest-Gang”, 2011); we provide here a scientific thread, leaving out the voluminous private correspondence among the principal players that Nickelsen and Govindjee (2011) examined in conjunction with their analysis of the principals’ published papers. We explore the development and course of the controversy and the ultimate resolution in favor of Emerson’s result as the phenomenon of the two-light-reaction, two-pigment-system scheme of photosynthesis came to be understood. In addition, we include a brief discussion of the discovery by Otto Warburg of the requirement for bicarbonate in the Hill reaction.     

The Influence of Light Wavelength on Phase-Dependent Responsiveness of the Photoperiodic Clock in Migratory Blackheaded Bunting

We investigated the phase-dependent effects of light wavelength on photoperiodic clock in the migratory blackheaded bunting. Two experiments were performed, employing a skeleton paradigm (6 hours light : 6 hours darkness : 1 hour light : 11 hours darkness; 6L : 6D : 1L : 11D) at 37 ± 2 lux intensity. In the experiment 1, both 6 and 1 h light pulses were given at the same wavelength, 500 nm (green) or 650 nm (red). A group exposed to both pulses of white light served as control. In the experi-ment 2, the two light pulses were given at two different wavelengths, 6 h at 500 nm (green) and 1 h at 640 nm (red) in one group or vice-versa in the other. There was almost no photoinduction when both light pulses in experiment 1, or 1 h light pulse in experiment 2, were green. On the other hand, birds fattened and testes recrudesced when both the light pulses in experiment 1, or 1 h light pulse in experiment 2, were red. Birds receiving both pulses of white light in experiment 1 showed an intermediate response. Taken together, these results indicate that the photoperiodic clock in buntings is differentially responsive at its various circadian phases to different light wavelengths.     

The Influence of Light Wavelength on Phase-Dependent Responsiveness of the Photoperiodic Clock in Migratory Blackheaded Bunting

We investigated the phase-dependent effects of light wavelength on photoperiodic clock in the migratory blackheaded bunting. Two experiments were performed, employing a skeleton paradigm (6 hours light : 6 hours darkness : 1 hour light : 11 hours darkness; 6L : 6D : 1L : 11D) at 37 ± 2 lux intensity. In the experiment 1, both 6 and 1 h light pulses were given at the same wavelength, 500 nm (green) or 650 nm (red). A group exposed to both pulses of white light served as control. In the experi-ment 2, the two light pulses were given at two different wavelengths, 6 h at 500 nm (green) and 1 h at 640 nm (red) in one group or vice-versa in the other. There was almost no photoinduction when both light pulses in experiment 1, or 1 h light pulse in experiment 2, were green. On the other hand, birds fattened and testes recrudesced when both the light pulses in experiment 1, or 1 h light pulse in experiment 2, were red. Birds receiving both pulses of white light in experiment 1 showed an intermediate response. Taken together, these results indicate that the photoperiodic clock in buntings is differentially responsive at its various circadian phases to different light wavelengths.     

Govindjee at 80: more than 50 years of free energy for photosynthesis

We provide here a glimpse of Govindjee and his pioneering contributions on the two light reactions and the two pigment systems, particularly on the water–plastoquinone oxido-reductase, Photosystem II. His focus has been on excitation energy transfer; primary photochemistry, and the role of bicarbonate in electron and proton transfer. His major tools have been kinetics and spectroscopy (absorption and fluorescence), and he has provided an understanding of both thermoluminescence and delayed light emission in plants and algae. He pioneered the use of lifetime of fluorescence measurements to study the phenomenon of photoprotection in plants and algae. He, however, is both a generalist and a specialist all at the same time. He communicates very effectively his passion for photosynthesis to the novice as well as professionals. He has been a prolific author, outstanding lecturer and an editor par excellence. He is the founder not only of the Historical Corner of Photosynthesis Research, but of the highly valued Series Advances in Photosynthesis and Respiration Including Bioenergy and Related Processes. He reaches out to young people by distributing Z-scheme posters, presenting Awards of books, and through tri-annual articles on “Photosynthesis Web Resources”. At home, at the University of Illinois at Urbana-Champaign, he has established student Awards for Excellence in Biological Sciences. On behalf of all his former graduate students and associates, I wish him a Happy 80th birthday. I have included here several tributes to Govindjee by his well-wishers. These write-ups express the high regard the photosynthesis community holds for “Gov” and illuminate the different facets of his life and associations.     

The influence of ATP/NADPH ratio on distribution of reductive pentose phosphate cycle metabolites as a possible causative factor of enhancement effect,chromatic transients and spectral changes of quantum yield of photosynthesis

A simple kinetic model for the reductive pentose phosphate cycle is suggested and analyzed. The changes in ATP/NADPH ratio caused by light wavelength alterations have been shown to result in the specific redistribution of metabolites in the cycle. The redistribution permits an explanation of the kinetic patterns of the enhancement effect and chromatic transients of photosynthesis. The spectra of relative ATP concentration have been calculated using enhancement values for green plants and cyanobacteria. Relative changes in quantum yield of photosynthesis were also calculated using data from spectra of relative ATP concentration changes. It is shown that changes in the quantum yield of photosynthesis may be fully explained by means of the spectral dependence of the relative ATP concentration in chloroplasts. It is concluded that the enhancement effect, chromatic transients and the ‘red drop’ of photosynthetic efficiency are not to be considered as exhaustive arguments for the Z-scheme of photosynthesis, although they do not contradict it.     

Evidence that putrescine modulates the higher plant photosynthetic proton circuit

The light reactions of photosynthesis store energy in the form of an electrochemical gradient of protons, or proton motive force (pmf), comprised of electrical (Δψ) and osmotic (ΔpH) components. Both components can drive the synthesis of ATP at the chloroplast ATP synthase, but the ΔpH component also plays a key role in regulating photosynthesis, down-regulating the efficiency of light capture by photosynthetic antennae via the q(E) mechanism, and governing electron transfer at the cytochrome b(6)f complex. Differential partitioning of pmf into ΔpH and Δψ has been observed under environmental stresses and proposed as a mechanism for fine-tuning photosynthetic regulation, but the mechanism of this tuning is unknown. We show here that putrescine can alter the partitioning of pmf both in vivo (in Arabidopsis mutant lines and in Nicotiana wild type) and in vitro, suggesting that the endogenous titer of weak bases such as putrescine represents an unrecognized mechanism for regulating photosynthetic responses to the environment.     

Mechanisms of amplification of photosynthetically active radiation in the symbiotic deep-water coral Leptoseris fragilis

The symbiotic coral Leptoseris fragilis lives in the Red Sea at depths of 95–145 m. Symbiotic dinoflagellates (zooxanthellae) themselves possess well known adaptations to low light intensities. In L. fragilis we found indications that light amplifying mechanisms of the host improve photosynthesis of the symbionts. Light of short wavelengths is absorbed by host pigments which transform short into longer wavelengths. The transformed light is more efficient for photosynthesis. Action spectra measurements of photosynthesis demonstrated the amplification of photosynthetically active radiation. Monochromatic light of 387 nm (outside the main absorption maxima of the algal pigments) at subsaturation photon flux densities was as effective photosynthetically as polychromatic light of 415–490 nm, which fits the absorption maxima of the zooxanthellae.     

Celebrating the millennium – historical highlights of photosynthesis research,Part 2

This paper is an introduction to Part 2 of our celebrations of the historical highlights of photosynthesis research. Part 1 was published in October 2002 as Volume 73 of Photosynthesis Research. After a brief introduction, we recognize two giants in the field: Cornelis B. van Niel (for anoxygenic photosynthesis), and Robert Hill (for oxygenic photosynthesis). This is followed by recognition of a 1960 book by Hans Gaffron, and a multi-authored book edited by W. Ruhland and André Pirson, and inclusion in the appendix of a list of selected books. Our celebration is enhanced by the inclusion of beautiful paintings of cells by Antoinette Ryter. After introducing all the historical papers contained in this volume, we honor Louis N. M. Duysens, one of the greatest biophysicists of our time, and finally we dedicate this volume to a great scientist, humanist and peacemaker: Eugene I. Rabinowitch. [12pt] 'Annihilating all that is made To a green thought in a green shade' – Andrew Marvell (1621–1678), The Garden (1681) This revised version was published online in August 2006 with corrections to the Cover Date.     

Verification of the Z-scheme in Chlamydomonas mutants with Photosystem I deficiency

In conflict with the Z-scheme of photosynthesis, it has recently been reported [Greenbaum et al. Nature (1995) 376: 438–441; Lee et al. Science (1996) 273: 364–367] that Photosystem II can drive ferredoxin reduction and photoautotrophic growth in some mutants of Chlamydomonas lacking detectable Photosystem I reaction centre, P700. Using the same mutants, B4 and F8, here we report that action spectra and parameters of flash yields of different photoreactions show the operation in ferredoxin-dependent H₂ photoproduction and CO₂ fixation of a fraction (at least 5% compared to wild- type) of the only Photosystem I complexes.