1: Biochemistry and metabolism
1. Photosynthesis in higher plants
• Autotrophic: simple mineral → complex organic compounds
• Photo-autotrophic: light energy → usable biochemical energy
Light-dependent reactions Carbon fixation reactions
In thylakoids In stroma
Oxidation H2O: H2O → O2 Reduction CO2: CO2 → carbohydrates
Produce ATP and NADPH Use ATP and NADPH
2. General concepts
• Absorb blue and red light (efficient in driving photosynthesis) → green light is reflected
• DIFFERENT PIGMENTS HAVE DIFFERENT ABSORPTION SPECTRA!
• Absorption photon → excited state (highly unstable)
1. Heat loss → lower excitation state
2. Re-emission photon (fluorescence) → ground state
3. Energy transfer to other molecules
4. Photo-chemical reaction
→must be fast, competition with other processes
• Chlorophylls (Chl a, Chl b, Bacteriochlorophyll a): absorb light
• Carotenoids (β-carotene): antenna pigment + photoprotective agent
• Bilin pigments (Phycoerythrobilin): antenna pigment + photoprotective agent
• Action spectra= wavelengths that stimulate most effectively PS
• Carotenoids transfer energy to chlorophylls → almost perfect overlap between absorption
and action spectra
3. Photosynthesis
• Happens in complexes containing light harvesting antennas (energy transfer) and
photochemical reaction centers (e- transfer)
➔ H+ gradient for ATP
production
PHOTOSYSTEM II (P680) PHOTOSYSTEM I (P700)
Absorption red light Absorption far-red light
Strong oxidant → oxidation water Strong reductant → reduction NADP+
• PSI and PSII: spatially separated in thylakoid membrane
1
, 1. PSI + LHCI dimer
2. PSII + LHCII trimer → in stacked regions
3. Cytochrome b6f
4. ATP synthase → uses H-gradient to produce ATP
• Diffusion of e- carriers between 2 PS:
1. Lipid-soluble organic redox co-factor plastoquinone = pool of reducing equivalents in
membrane
2. Cu-containing protein plastocyanin in thylakoid lumen
4. Organisation of light-absorbing antenna systems
• Evolutionary adaptation to different environments
• Large membrane-associated pigment-protein complexes
• Very efficient FRET
• Energy is trapped and funneled to reaction centers: carotenoids → chlorophyll b →
chlorophyll a → P680* (red shift)
5. Mechanisms of electrontransport
• E- travel from H2O → NADP+ through different
electron carriers with increasing redox potential
• E- transfer: associated with H+ transfer → drives
ATP synthesis
1. PSII oxidizes water → produces H+ in lumen
2. Cytochrome b6f oxidizes PQH2 (reduced by
PSII), coupled with H+ transfer
3. PSI reduces NADP+ via ferredoxin (Fd) and
Fd-NADP+ reductase (FNR) → NADPH
4. Using H+ gradient and H+ back-diffusion ATP synthase → ATP
• Energy is captured when excited chlorophyll reduces an e- acceptor molecule
• Plastoquinone accepts e- from PSII:
1. Pheophytin is early intermediate acceptor
2. E- then passed on to complex of 2 plastoquinones → reduction: 2 e- for each PQ
Electron flow through Cyt b6f complex also transports H+
• E- and H+ flow according to Q-cycle: 1 e- to PC-PSI + 1 e-
to Q → 2 H+ to lumen
1. First QH2 hydrolysed to Q (loses 2 e- and 2 H+) →
release of 2 H+ in lumen
2. 1 e- → plastocyanin → PSI
3. 1 e- → cycled to oxidized quinone
4. Second QH2 is oxidized → release of 2 H+ in lumen
5. 1 e- → plastocyanin → PSI
6. 1 e- + 2H+ from stroma to Q-radical → QH2
→4 H+ transported in lumen for each 2e- delivered to PSI
Two e- flow trajectories from PSI:
1) Reduction of NADP+ via Fd and FNR → NADPH
2) Cyclic e- transport (back to cyt b6f) → H+ gradient → ATP
Herbicides that block photosynthetic e- flow:
2
, • DCMU: competes for PQ binding site at PSII
• Paraquat: accepts e- from PSI + passes it to O2 → 02- superoxide (damage chloroplast lipids)
6. Proton transport and ATP-synthesis
• Photo-phosphorylation= light-dependent ATP production
• Chemi-osmotic mechanism:
- Similar to aerobic respiration in bacteria and mitochondria (ATP synthesis) and transport
of many ions and metabolites (conversely using ATPases)
- Chemical potential (H+ gradient, acidification lumen)= source of energy
→only H+ gradient needed for ATP (without light)
• Proton motive force drives ATP synthase complex in stroma lamellae + at edges grana stacks
7. Repair and regulation of photosynthetic machinery
• Designed to absorb large amounts of light energy → excess
energy is damaging because of production of toxic ROS
Carotenoids also serve as photoprotective agents:
• Chlorophyll in excited state can react with molecular oxygen →
ROS
• Excited state of carotenoids does not have enough energy to
form singlet oxygen → decays to ground state while losing E as
heat
→zeaxanthin, violaxanthin, antheraxanthin
• PSI is vulnerable to damage by ROS:
- Ferredoxin: strong reductant → reduce molecular oxygen to
superoxide → eliminated by superoxide dismutase +
ascorbate peroxidase
2: Photosynthesis, the C
reactions
1.The Calvin Benson cycle
1. Carboxylation of CO2-acceptor: ribulose-1,5-biP + CO2+ H2O →
3-phosphoglycerate
• RUBISCO
2. Reduction of 3-phosphoglycerate: 3-phosphoglycerate + ATP +
NADPH → glyceraldehyde-3P
3. Regeneration of CO2-acceptor: glyceraldehyde-3P + ATP →
ribulose-1,5-biP
• Induction period= build-up concentration intermediates +
light activation of enzymes!!!
• RUBISCO: high affinity for O2 → evolved in oxygen-poor
environment
3
, • Carboxylase: 3-phosphoglycerate
• Oxygenase: 3-phosphoglycerate + 2-phosphoglycolate
• 3-phosphoglycerate is enzyme inhibitor → plants found way to deal with this!
IMPORTANT ENZYMES IN CYCLE:
Ribulose-1,5-biP Ribulose-1,5-biP + CO2 + H2O → 2 x 3-phosphoglycerate
carboxylase/oxygenase (RUBISCO)
3-phosphoglycerate kinase 3-phosphoglycerate + ATP → 1,3-biphosphoglycerate + ADP
NADP-glyceraldehyde-3P 1,3-biphosphoglycerate + NADPH + H+ → glyceraldehyde-3P
dehydrogenase + NADP+ + Pi
Phosphoribulokinase Ribulose-5P + ATP → ribulose-1,5-biP + ADP+H+
2.Regulation of Calvin-Benson cycle
A. Enzyme levels:
- Gene expression (transcription), protein biosynthesis (translation) and stability
- Coordination via retrograde signaling (plastids → nucleus) + anterograde signaling
(nucleus → plastids)
B. Specific activity:
- Posttranslational modifications!
- Changes in covalent bonds
- Modification of non-covalent interactions by changes in ionic composition cellular
environment, binding of enzyme effectors, association with regulatory proteins in supra-
molecular complexes, interaction with thylakoid membranes
1. RUBISCO-ACTIVASE REGULATES THE CATALYTIC ACTIVITY OF RUBISCO
Rubisco activase (ATPase):
Removes/ lowers affinity for sugar-Ps that prevent activation of inactive
rubisco or inhibit catalysis of active rubisco
→ redox-regulated when reduced (thiol): affinity for ATP rises
CO2 as activator:
-carbamylation and metal ion binding
-higher pH and [Mg2+] → stimulate activation after illumination
2. LIGHT REGULATES THE CALVIN-BENSON CYCLE VIA FERREDOXIN-
THIOREDOXIN SYSTEM (REDUCTION)
3. LIGHT-DEPENDENT ION MOVEMENTS MODULATE CYCLE ENZYMES
Light → H+ in thylakoid lumen → increased pH + coupled release Mg2+ in stroma →
stimulates activity of rubisco
4
1. Photosynthesis in higher plants
• Autotrophic: simple mineral → complex organic compounds
• Photo-autotrophic: light energy → usable biochemical energy
Light-dependent reactions Carbon fixation reactions
In thylakoids In stroma
Oxidation H2O: H2O → O2 Reduction CO2: CO2 → carbohydrates
Produce ATP and NADPH Use ATP and NADPH
2. General concepts
• Absorb blue and red light (efficient in driving photosynthesis) → green light is reflected
• DIFFERENT PIGMENTS HAVE DIFFERENT ABSORPTION SPECTRA!
• Absorption photon → excited state (highly unstable)
1. Heat loss → lower excitation state
2. Re-emission photon (fluorescence) → ground state
3. Energy transfer to other molecules
4. Photo-chemical reaction
→must be fast, competition with other processes
• Chlorophylls (Chl a, Chl b, Bacteriochlorophyll a): absorb light
• Carotenoids (β-carotene): antenna pigment + photoprotective agent
• Bilin pigments (Phycoerythrobilin): antenna pigment + photoprotective agent
• Action spectra= wavelengths that stimulate most effectively PS
• Carotenoids transfer energy to chlorophylls → almost perfect overlap between absorption
and action spectra
3. Photosynthesis
• Happens in complexes containing light harvesting antennas (energy transfer) and
photochemical reaction centers (e- transfer)
➔ H+ gradient for ATP
production
PHOTOSYSTEM II (P680) PHOTOSYSTEM I (P700)
Absorption red light Absorption far-red light
Strong oxidant → oxidation water Strong reductant → reduction NADP+
• PSI and PSII: spatially separated in thylakoid membrane
1
, 1. PSI + LHCI dimer
2. PSII + LHCII trimer → in stacked regions
3. Cytochrome b6f
4. ATP synthase → uses H-gradient to produce ATP
• Diffusion of e- carriers between 2 PS:
1. Lipid-soluble organic redox co-factor plastoquinone = pool of reducing equivalents in
membrane
2. Cu-containing protein plastocyanin in thylakoid lumen
4. Organisation of light-absorbing antenna systems
• Evolutionary adaptation to different environments
• Large membrane-associated pigment-protein complexes
• Very efficient FRET
• Energy is trapped and funneled to reaction centers: carotenoids → chlorophyll b →
chlorophyll a → P680* (red shift)
5. Mechanisms of electrontransport
• E- travel from H2O → NADP+ through different
electron carriers with increasing redox potential
• E- transfer: associated with H+ transfer → drives
ATP synthesis
1. PSII oxidizes water → produces H+ in lumen
2. Cytochrome b6f oxidizes PQH2 (reduced by
PSII), coupled with H+ transfer
3. PSI reduces NADP+ via ferredoxin (Fd) and
Fd-NADP+ reductase (FNR) → NADPH
4. Using H+ gradient and H+ back-diffusion ATP synthase → ATP
• Energy is captured when excited chlorophyll reduces an e- acceptor molecule
• Plastoquinone accepts e- from PSII:
1. Pheophytin is early intermediate acceptor
2. E- then passed on to complex of 2 plastoquinones → reduction: 2 e- for each PQ
Electron flow through Cyt b6f complex also transports H+
• E- and H+ flow according to Q-cycle: 1 e- to PC-PSI + 1 e-
to Q → 2 H+ to lumen
1. First QH2 hydrolysed to Q (loses 2 e- and 2 H+) →
release of 2 H+ in lumen
2. 1 e- → plastocyanin → PSI
3. 1 e- → cycled to oxidized quinone
4. Second QH2 is oxidized → release of 2 H+ in lumen
5. 1 e- → plastocyanin → PSI
6. 1 e- + 2H+ from stroma to Q-radical → QH2
→4 H+ transported in lumen for each 2e- delivered to PSI
Two e- flow trajectories from PSI:
1) Reduction of NADP+ via Fd and FNR → NADPH
2) Cyclic e- transport (back to cyt b6f) → H+ gradient → ATP
Herbicides that block photosynthetic e- flow:
2
, • DCMU: competes for PQ binding site at PSII
• Paraquat: accepts e- from PSI + passes it to O2 → 02- superoxide (damage chloroplast lipids)
6. Proton transport and ATP-synthesis
• Photo-phosphorylation= light-dependent ATP production
• Chemi-osmotic mechanism:
- Similar to aerobic respiration in bacteria and mitochondria (ATP synthesis) and transport
of many ions and metabolites (conversely using ATPases)
- Chemical potential (H+ gradient, acidification lumen)= source of energy
→only H+ gradient needed for ATP (without light)
• Proton motive force drives ATP synthase complex in stroma lamellae + at edges grana stacks
7. Repair and regulation of photosynthetic machinery
• Designed to absorb large amounts of light energy → excess
energy is damaging because of production of toxic ROS
Carotenoids also serve as photoprotective agents:
• Chlorophyll in excited state can react with molecular oxygen →
ROS
• Excited state of carotenoids does not have enough energy to
form singlet oxygen → decays to ground state while losing E as
heat
→zeaxanthin, violaxanthin, antheraxanthin
• PSI is vulnerable to damage by ROS:
- Ferredoxin: strong reductant → reduce molecular oxygen to
superoxide → eliminated by superoxide dismutase +
ascorbate peroxidase
2: Photosynthesis, the C
reactions
1.The Calvin Benson cycle
1. Carboxylation of CO2-acceptor: ribulose-1,5-biP + CO2+ H2O →
3-phosphoglycerate
• RUBISCO
2. Reduction of 3-phosphoglycerate: 3-phosphoglycerate + ATP +
NADPH → glyceraldehyde-3P
3. Regeneration of CO2-acceptor: glyceraldehyde-3P + ATP →
ribulose-1,5-biP
• Induction period= build-up concentration intermediates +
light activation of enzymes!!!
• RUBISCO: high affinity for O2 → evolved in oxygen-poor
environment
3
, • Carboxylase: 3-phosphoglycerate
• Oxygenase: 3-phosphoglycerate + 2-phosphoglycolate
• 3-phosphoglycerate is enzyme inhibitor → plants found way to deal with this!
IMPORTANT ENZYMES IN CYCLE:
Ribulose-1,5-biP Ribulose-1,5-biP + CO2 + H2O → 2 x 3-phosphoglycerate
carboxylase/oxygenase (RUBISCO)
3-phosphoglycerate kinase 3-phosphoglycerate + ATP → 1,3-biphosphoglycerate + ADP
NADP-glyceraldehyde-3P 1,3-biphosphoglycerate + NADPH + H+ → glyceraldehyde-3P
dehydrogenase + NADP+ + Pi
Phosphoribulokinase Ribulose-5P + ATP → ribulose-1,5-biP + ADP+H+
2.Regulation of Calvin-Benson cycle
A. Enzyme levels:
- Gene expression (transcription), protein biosynthesis (translation) and stability
- Coordination via retrograde signaling (plastids → nucleus) + anterograde signaling
(nucleus → plastids)
B. Specific activity:
- Posttranslational modifications!
- Changes in covalent bonds
- Modification of non-covalent interactions by changes in ionic composition cellular
environment, binding of enzyme effectors, association with regulatory proteins in supra-
molecular complexes, interaction with thylakoid membranes
1. RUBISCO-ACTIVASE REGULATES THE CATALYTIC ACTIVITY OF RUBISCO
Rubisco activase (ATPase):
Removes/ lowers affinity for sugar-Ps that prevent activation of inactive
rubisco or inhibit catalysis of active rubisco
→ redox-regulated when reduced (thiol): affinity for ATP rises
CO2 as activator:
-carbamylation and metal ion binding
-higher pH and [Mg2+] → stimulate activation after illumination
2. LIGHT REGULATES THE CALVIN-BENSON CYCLE VIA FERREDOXIN-
THIOREDOXIN SYSTEM (REDUCTION)
3. LIGHT-DEPENDENT ION MOVEMENTS MODULATE CYCLE ENZYMES
Light → H+ in thylakoid lumen → increased pH + coupled release Mg2+ in stroma →
stimulates activity of rubisco
4
