1. §4 Mechanism of Photosynthesis

2. Photosynthesis comprises two steps: Light reaction and Dark reaction
Warburg found photosynthesis can be divided into two steps in 1920
Light reaction
Photosynthesis
Dark reaction

3. Flash experiment with Chlorella (小球藻)
Radiated continuously
Less O2
Light
Radiated for S , interval S, 50cps
More O2

4. Now it’s clear Light reaction: Dark reaction:
Primary reaction: absorption, transfer and conversion of light energy to electric energy.
Electron transfer and photophosphorylation: turn electric energy into active chemical energy.
Dark reaction:
Assimilation of CO2, turn active chemical energy to stable chemical energy.

5. I: Primary reaction The first step of photosynthesis
Includes: absorption, transference and conversion of light energy
Site: thylakoid

6. Photosynthetic Unit
Photosynthetic unit: the minimum unit for carrying out photosynthetic O2 production
Emerson & Arnold (1930) found that it took the cooperative action of around 2400 chlorophyll molecules to produce one molecule of O2.
It takes a minimum of 8 photons to produce one molecule of O2.
For each photon, about 300 chls are needed.

7. A Photosynthetic Unit includes:
Light harvesting system: includes antenna pigments and proteins (chl bond with proteins), light harvest and transmission
Reaction center: converse light energy to electric energy. It includes:
Reaction center chl: some special chla
Primary electron donor
Primary electron acceptor
And related proteins

9. How energy is converted?
light
DPA → DP*A →DP+A- →D+PA-

10. Note: resonance energy transfer is quite rapid
Note: resonance energy transfer is quite rapid. The energy of 1 red photon can be passed to chl (30ps/s) in its life span. The efficiency is up to %

12. Antenna complex as energy funnel

13. II: Electron transfer and Photophosphorylation
The electron produced in the primary reaction is transferred through the photosynthetic electron transfer chain, and at last causes the generation of ATP and NADPH.

14. 1. Electron transference needs two photosystems
PSI and PSII
Discovered by Emerson from his famous Red Drop Phenomenon (红降现象）and Enhancement Effect（双光增益效应）.

15. 685 nm

16. 685nm
650nm
650nm
685nm

17. Conclusion:
Light reaction needs 2 photosystems, one is driven by long wavelength light, another is driven by short wavelength light.

18. 2 PSII, PSI and the chain

19. A. Four protein complexes in the thylakoid membrane

20. 1) PSII Components P680 Primary acceptor: pheo Primary donor: LHCII
Peptides(D1and D2;CP43,CP47,cytb6/f)
QA，QB
OEC(33,23 and 16kDa)
Functions:
photolysis of H2O to emit O2 and provide e and proton

21. Events at the PS II reaction center

23. Hill Reation
The Hill reaction occurs when isolated ‘chloroplasts’ are illuminated in the presence of an electron acceptor ‘A.’ The acceptor is reduced (to AH2) and molecular oxygen (O2) is evolved. A can be NAD(P)+ and so on.
2H2O+2A　　　　　2AH2+O2
light
chloroplast

24. Significance of the discovery
Proved that O2 is released from H2O, rather than CO2, as had been previously thought. 
Proved that chloroplast play an important role in photosynthesis.
Proved that photosynthesis is an oxidation-reduction reaction, and the process of photosynthesis is the transport of activated electron from one substance to another. This established the essence of converting light  energy to chemical energy.

25. Proof of water oxidation clock (Joliet light flash experiment:5-10s, interval,300ms)
Note: maximum O2 evolution comes after the 3rd flash, because during dark relaxation, most OEC returns to S1 state.

26. How O2 is evolved? ----the “water oxidation clock”
OEC contains 4 Mn, thus shows 5 oxidative states: S0，S1，S2，S3，S4. : S0 and S1 are stable states and in the dark, S0 is ¼ and S1 is 3/4， S2，S3 are unstable and converted to S0 and S1 in the dark. For each time P680 absorbs a photon, P680 captures a electron form OEC through Tyr and OEC accumulates a positive charge. Once OEC accumulates 4 charges, it will oxidate H2O and split it to evolve O2.

27. Structure of Mn-cluster of OEC
Note: there are also Cl, O and Ca in this model

28. 2) PSI Components Function：reduce NADP+ to NADPH P700
Primary electron acceptor：A0
Primary electron donor：PC
Electron transporter
LHCI
Peptites(PsaA:82kDa),Psa:83kDa; C,D,E,F…)
Function：reduce NADP+ to NADPH

29. 3) cytb6/f complex Components:
4 peptides, of which 3 contain Fe (1 cytb6, 1 cytf, 1 Rieske Fe)
Function:
Electron transport
PQ shuttle

30. 4) ATP Synthase Components: CF0：I II III IV(1:1:12:1)
Function:
Generate ATP

31. B: Electron Transfer Chain

34. Electron transport types:
①Noncyclic electron transport, ATP and NADPH;
②cyclic electron transport, only ATP;
③pseudocyclic electron transport, (ATP and ROS) ③
O2-
O2- ① O2 ②

35. Inhibitors of Electron Transfer Chain
PSII cytb6/f PSI
DCMU(敌草隆)：QA→QB
DBMIB(2,5-二溴-3-甲基-6-异丙基-对-苯醌)：PQ oxidation
Paraquat(百草枯)：deviate from FA/FB

36. C: Proton accumulation in the lumen
Two sources:
(1) Photolysis of water
(2) PQ shuttle

38. D: Photophosphorylation
Equipment: ATP synthase=CF
Driving force: ΔμH+ across the thylakoid membrane
Mechanism:
① chemiosmotic theory of Michell.
② binding change theory of Boyer

39. Chemiosmotic theory---- Transmembrane H+ gradient generates ATP

40. Proved by Jagenhorf

41. Binding Change Mechanism

42. Inhibitors of Photophosphorylation
Uncoupler: DNP(二硝基苯酚) , FCCP(羰基-氰-对-三氟甲氧基苯腙）
ATPase inhibitor: oligomycin（寡霉素）, prevent H+ to across CF0
Inhibitor of the chain: DCMU, DBMIM paraquat

43. Summary of Light Reaction
To evolve 1 molecule O2, 4 electrons are needed, thus theoretically 8 photons are required to evolve 1 O2.
2H2O → 4H+　＋4e + O2
During light reaction, O2 is evolved together with the production of ATP and NADPH, which will be consumed in dark reaction for CO2 assimilation, thus ATP and NADPH are named assimilation power. -2