1. Photosynthesis: Overview of photosynthesis, conversion of light energy to redox energy, and the Z scheme Bioc 460 Spring 2008 - Lecture 31 (Miesfeld) Solar power can provide a sustainable energy source for some areas in the world Paraquat is an inhibitor of PSI that was used to kill marijuana plants Agricultural crops are the primary means for converting solar energy into chemical energy for life on earth Paraquat

2. The energy from sunlight is used to initiate photooxidation reactions in light-absorbing pigments that convert light energy into chemical energy. The light reactions of photosynthesis are similar to the electron transport system in that they require a proton impermeable membrane and a series of linked redox reactions to generate proton motive force used for ATP synthesis. Photooxidation uses the oxidation of H 2 O to produce O 2 in a process that provides electrons for photophosphorylation and the reduction of NADP+ to produce NADPH. Chemical energy in the form of ATP and NADPH is used to convert CO 2 to glyceraldehyde-3-P using enzymes in the Calvin Cycle pathway. Plants use sunlight during the day and aerobic respiration at night. Key Concepts in Photosynthesis

3. The photosynthetic electron transport system is often referred to as the light reactions of photosynthesis, whereas, the Calvin cycle has been called the dark reactions. However, the term "dark reactions" can be misleading because the Calvin cycle is most active in the light when ATP and NADPH levels are high.

4. Net reaction of photosynthesis and carbon fixation The O 2 generation is the result of H 2 O oxidation, whereas, the CO 2 is used to synthesize carbohydrate (CH 2 O) It takes 2 H 2 O to make an O 2 and six CO 2 molecules are required for the synthesis of each molecule of glucose, therefore: ΔGº' for this reaction is +2868 kJ/mol! This is overcome by the energy potential stored in the products of photosynthetic electron transport, namely, ATP and NADPH. H 2 O + CO 2 ─ (light energy) → (CH 2 O) + O 2 6 H 2 O + 6 CO 2 ─ (light energy)→ C 6 H 12 O 6 + 6 O 2

5. The Biosphere Experiment, then and now! Biosphere 2 did not actually work very well because CO 2 levels built up inside the sealed environment and periodic CO 2 removal was required. The first “Biospherian” came out after ~1 year. Joseph Priestly - 1772 Biosphere 2 - 1991

6. 1. What do the photosynthetic electron transport system and Calvin cycle accomplish for the cell? The photosynthetic electron transport system converts light energy into redox energy which is used to generate ATP by chemiosmosis and reduce NADP + to form NADPH. Calvin cycle enzymes use energy available from ATP and NADPH to reduce CO 2 to form glyceraldehyde-3-P, a three carbon carbohydrate used to synthesize glucose. Photosynthetic cells use the carbohydrate produced by the Calvin cycle as a chemical energy source for mitochondrial respiration in the dark. Photosynthetic organisms are autotrophs because they derive energy from light rather than from organic materials (as food).

7. 2. What are the overall net reactions of photosynthetic electron transport system and the Calvin cycle? Photosynthetic electron transport system (production of ATP and O 2 ): 2 H 2 O + 8 photons + 2 NADP + + ~3 ADP + ~3 Pi → O 2 + 2 NADPH + ~3 ATP Calvin cycle (six turns of cycle yields glucose): 6 CO 2 + 12 NADPH + 18 ATP + 12 H 2 O → Glucose + 12 NADP + + 18 ADP + 18 Pi

8. 3. What are the key enzymes in the photosynthetic electron transport system and the Calvin cycle? Protein components of the photosynthetic electron transport system – three protein complexes are required for the oxidation of H 2 O and reduction of NADP + ; photosystem II (P 680 reaction center), cytochrome b 6 f (proton pump) and photosystem I (P 700 reaction center). Chloroplast ATP synthase – enzyme responsible for the process of photophosphorylation which converts proton-motive force into net ATP synthesis; this enzyme is very similar to mitochondrial ATP synthase. Ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) - is responsible for CO 2 fixation in the first step of the Calvin cycle. Rubisco activity is maximal in the light when stromal pH is ~8 and Mg 2+ levels are elevated due to proton pumping.

9. 4. What are examples of the photosynthetic electron transport system and Calvin Cycle in real life? DCMU (dichlorophenyl dimethylurea) is a broad spectrum herbicide that functions by blocking electron flow through photosystem II and is used to reduce weeds in non-crop areas. Another herbicide, paraquat, prevents reduction of NADP + by accepting electrons from intermediate reductants in photosystem I. PSII PSI

10. Overview of Photosynthesis Light energy is used to oxidize 2 H 2 O which releases 4e- and 4H+ The photosynthetic electron transport system and photophosphorylation work togethe to generate ATP and NADPH for sugar synthesis by the Calvin Cycle 4e- Redox reactions are used to translocate an additional 8H+

11. What pathway has GAP as a central intermediate, does this make sense here? Pay attention to the compartmentalization inside and outside of the chloroplast. The product of the carbon fixation is glyceraldehyde-3P (GAP) which is converted to hexose sugars for use as chemical energy at night.

12. Peter Mitchell’s chemiosmotic theory was actually first proven using photosynthetic systems in which an artificial proton gradient was established using buffered solutions at different pH values. What would happen to the ATP* if the buffer was now switched back to pH4? 1 2 3 4 5

13. The chloroplast ATP synthase is structurally and functionally similar to the mitochondrial ATP synthase except that 4 H + are required for every ATP synthesized based on experiments showing that ~3 ATP are synthesized for every O 2 generated (12 H + transported/O 2 generated).

14. Stroma thylakoids Granal thylakoids Outer envelope Inner envelope Chloroplast Structure

15. Light energy is absorbed by numerous accessory pigments which can transfer the absorbed energy to nearby reaction centers containing specialized chlorophyll molecules. These accessory pigments function as light harvesting antenna.

16. Chlorophylls are the primary light gathering pigments They have a heterocyclic ring system that constitutes an extended polyene structure, which typically has strong absorption in visible light.

17. Plant pigments cover a broad absorption spectra

18. Organization of Photosynthetic pigments Light absorbing pigments are organized in functional arrays called photosystems of which there are two; PSII and PSI in plants. Light harvesting or antenna pigment molecules are specialized to absorb light and transfer the energy to neighboring pigment molecules. Photochemical reaction center pigment molecules are specialized to transduce light energy into chemical energy. Several hundred light harvesting pigment molecules funnel light to one reaction center molecule.

19. Light harvesting pigment molecules transfer energy to nearby molecules which eventually results in photooxidation Top view of antenna chlorophylls. Note that both PSI and PSII are reaction centers.

20. Z Scheme of Photosynthetic Electron Transport Light energy absorbance “kicks” electrons into higher energy states which is used to convert solar energy into redox energy when the electrons are passed one at a time to electron carriers with increasingly positive standard reduction potentials. The redox energy is captured in the form of chemical energy (ATP, NADPH.

21. The oxidation of 2 H 2 O to produce one mole of O 2 requires the absorbance of 8 photons to move the 4 e- through the entire photosynthetic electron transport system.

22. Energy input at both the PSII and PSI reaction centers

23. Light harvesting involves resonance energy transfer, whereas, photooxidation involves e- transfer from chlorophyll to the electron acceptor called pheophytin which becomes negatively charge as denoted by Pheo-. Importantly, the oxidized chlorophyll molecule (now positively charged, Chl + ) returns to the ground state by accepting an electron through a coupled redox reaction involving the oxidation of H 2 O. Light absorption kicks an e- into a higher orbital, what happens next is either energy transfer, photooxidation, or fluorescence.

24. Now all we have to do is follow the photons, electrons, and protons starting with PSII

25. Photosystem II (PSII) Photosystem II contains chlorophylls a and b and absorbs light at 680nm. This is a large protein complex that is located in the thylakoid membrane.

26. Functional organization of the PSII complex

27. The electron that was transferred from the P680 chlorophyll reaction center needs to be replaced, this replacement electron comes from the oxidation of H 2 O within the oxygen evolving complex. The tricky part is that the oxidation of H 2 O releases 4 e-, however, photooxidation only transfers one e- at a time to pheophytin. Therefore, the Mn atoms must be able to “store” electrons and release them one at a time.

28. Functional organization of the cytochrome b 6 f complex The same deal here as in complex III of ETS, we need to convert the 2 e- carrier PQBH 2 into a 1 e- carrier in PC. The answer is the Q cycle, duh!

29. The light-driven Q cycle is responsible for translocation of 8 H + from the stroma to the lumen ATP synthase complex

30. Photosystem I (PSI) The final stage of photosynthesis: the absorption of light energy by PS I is at a maximum of 700 nm. Again 4 photons are absorbed, but in this case, the energy is used to generate reduced ferredoxin, which is a powerful reductant. Structure of PS I complex showing Fe-S clusters

31. Functional organization of the PSI complex

32. Ferrodoxin NADP + reductase plays a crucial role in converting redox energy into a useable form for the Calvin Cycle by generating NADPH. Since e- arrive in PSI one at a time, the FAD coenzyme must store on e- in a semiquinone chemical structure.

33. Paraquat was once used extensively as an aerial herbicide to destroy illegal fields of marijuana and coca plants in North and South America. However its use was discontinued because smoking paraquat-contaminated plants causes lung damage.