Biology Chapter 18 Summary
Chapter 18
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Takusagawa’s Note©
Chapter 18: Photosynthesis
1. CHLOROPLASTS - Photosynthesis is carried out at chloroplasts. Structure of chloroplast
Outer membrane Stroma lamellae Inner membrane Thylakoid
Chloroplast
Granum Dark reaction Stroma Light reaction
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Composition of innermembrane and granum membrane are unusual. - Phospholipid (negatively charged) ~10% - Neutral lipid (galactose) ~80%
Photosynthesis occurs in two distinct phases: 1. Light reactions --- Generates NADPH & ATP by using light energy at thylakoid membrane (prokaryotes: inner membrane). 2. Dark reactions --- Synthesize carbohydrates from CO2 + H2O by using NADPH & ATP at stroma.
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LIGHT REACTION Initially believed: CO2 + H2O ⎯light → CH2O + O2 ⎯ van Niel showed: CO2 + 2H2S ⎯light → CH2O + 2S + H2O ⎯ In general: CO2 + 2H2A ⎯light → CH2O + 2A + H2O ⎯
This suggested that photosynthesis is two step reactions: 1. Light energy oxidizes H2A (light reaction): 2H2A ⎯light → 2A + 4[H] ⎯ 2. The reducing agent [H] reduces CO2 (dark reaction): 4[H] + CO2 ⎯→ CH2O + H2O
A. Absorption of light - The principal photoreceptor is chlorophyll (Chl) derived from protoporphyrin IX. - Major differences between chlorophyll and hemes are: - Chl has an extra ring (V) (cyclopentanone). - Ring IV of Chl is more reduced. - …show more content…
Mg is present instead of Fe. - Side chains are different --- some Chl have a long chain at R4 site.
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Chapter 18 Eukaryotes have: - Chlorophyll a (Chl a) - Chlorophyll b (Chl b) Prokaryotes have: - Bacteriochlorophyll a (BChl a) - Bacteriochlorophyll b (BChl b)
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- Note: No absorption of Chl a and b in 480 - 620 nm region. But other pigments, carotenoids (such as β-carotene), phycoerythrin and phycocyanin absorb the light between 480 and 620.
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Chapter 18 Photo-energy Photo-energy E = hν = hc
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Takusagawa’s Note©
light (2.998 × 108 m/s), and λ = wavelength of light. - One mole of red light (6.02 × 1023 photons = 1 einstein) with λ= 700 nm has 171 kJ/einstein: Nhc (6 × 10 23 )(6.6 × 10 −34 J ⋅ s −1 )(3 × 10 8 m ⋅ s −1 ) E= = = 171kJ / einstein . λ 700 × 10 −9 m - In general, when molecules absorb photon energy, they move from ground state to various excited states. - There are four pathways that an excited state returns to lower energy state. 1. Losing energy as heat (kinetic). 2. Losing energy as light (fluorescence) that normally shifts to longer wavelength. 3*. Losing energy as energy transfer to another molecule. ⎯ MA + MB ⎯light → M*A + MB ⎯→ MA + M*B * 4 . Losing energy by losing electron (photooxidation) ⎯ Chl ⎯light → Chl+ + e* 3 and 4 are important for photosynthesis.
λ
, where h = Planck’s constant (6.626 × 10-34 J/s), c = speed of
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Light absorbed by antenna chlorophylls and accessory pigments is transferred to photosynthetic reaction centers - Light is absorbed in “light-harvesting complexes” (LHCs) located on the membrane of thylakoid in chloroplast. - LHC is membrane bound proteins containing Chl molecules and other pigments.
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Light-harvesting complex in pea chloroplast Most Chl molecules and other pigment molecules absorb light and the light energy (exciton) is randomly transferred to the neighboring Chl or other pigment until it is trapped by a “photosynthetic reaction center”. Thus, the function of most Chl’s is to gather light, i.e., they act as light-harvesting antennas. Photosynthetic reaction centers contain the same Chl molecules as antennas have. However, Chl molecules in photosynthetic reaction centers have slightly lower excited state energies because their different environments. Thus, excitons are trapped or funneled into reaction centers. A photosynthetic reaction center is surrounded by ~300 photosynthetic antenna complexes. Photosynthetic reaction center
Schemetic diagram of light harvesting complex
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Chloroplast
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C. Plant Photosynthesis is two-center electron transport - Bacteria photosynthesis is one center process, i.e., one light absorbing event, whereas - Photosynthesis in plants and algae is two center process, i.e., two light absorbing event. light (4 photons) 1. 2H2O ⎯⎯⎯⎯⎯⎯⎯ ⎯→ O2 + 4e- + 4H+ light (4 photons) ⎯ 2. 2NADP+ + 4e- + 2H+ ⎯ ⎯⎯⎯⎯⎯⎯ → 2NADPH Thus, light is used to reduce NADP+ with H2O. 2NADP+ + 2H2O ⎯→ 2NADPH + O2 + 2H+
ΔE°′ = -0.815 V ΔE°′ = -0.320 V ΔE°’ = -1.135 V
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ΔG°’ = -nFΔE°’ = 438 kJ/mol, where n = 4 (since 4 electrons are involved) and F=96.5 kJ/V.
As explained in previously, photo-energy of 700 nm light per one mole of photon (einstein) is: E = 171 kJ/einstein. In each event, four electrons are involved. Thus the total electrons are eight (Remember: two-light absorbing events). One electron activation requires one photon. Thus eight photons are necessary to activate eight electrons. The input light energy is: E = 8 × 171 = 1368 kJ/mol, which is much larger than ΔG°’(438 kJ/mol) of 2 mol NADPH and 1 mol O2 productions.
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ΔG°’ can be calculated from the pair of half-cell reactions listed in Table 15-4. NADP+ + H+ + 2e- → NADPH E°’ = -0.320 V + H2O → ½O2 + 2H + 2e E°’ = -0.815 V Thus, 2 mol NADPH and 1 mol O2 production require 2 × (-0.320 + -0.815) = -2.27 V. Since ΔG°’ = -nFΔE°’, ΔG°’ = -2 × 96.5 × -2.27 = 438.11 kJ/mol
Photosynthetic O2 production requires two-sequential photosystems - These events are occurred in two different photosynthetic reaction centers that are connected essentially in series. 1. Photosystem II (PSII) --- generates a weak reductant and strong oxidant to oxidize H2O. light (4 photons) 2H2O ⎯⎯⎯⎯⎯⎯⎯ → O2 + 4e- + 4H+ ⎯
eChl light Chl* Chl+ e- (from H2O)
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2. Photosystem I (PSI) --- generates a strong reductant to reduce NADP+ and weak oxidant. light (4 photons) 2NADP+ + 4e- + 2H+ ⎯ ⎯⎯⎯⎯⎯⎯ → 2NADPH ⎯
Chl
light
eChl* Chl-
(to NADP+) e
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Herbicide dichlorophenyl-dimethylurea (DCMU) blocks photosynthetic electron transport from PSII to cytochrome b6-f.
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Three complexes in membrane - are involved in the electron transport from (2H2O → O2 + 4H+ + 4e-) event to (2NADP+ + 2H+ + 4e- → 2NADPH) event. Those are: 3) PSI 1) PSII 2) Cyt b6-f complex - The electrons are carried by the mobile electron carriers between the complexes. Plastoquinol (PQ) --- between PSII and Cyt b6-f complex. Plastocyanin (PC) ---between Cyt b6-f complex and PSI. - Electron flow: PSII → (PQ) → Cyt b6-f complex → (PC) → PSI
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Plastoquinol (PQ) has two oxidation states, PQ (oxidized state) and PQH2 (reduced state). Plastocyanin (PC) is a small Cu-containing protein.
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Details of electron transport 1. Oxidation of H2O by “O2-evolving complex” (OEC) in PSII. - OEC contains 4Mn cluster that binds 2H2O. - Oxidation of H2O occurs 5 steps, i.e., S0, S1, S2, S3, S4. Four electrons are stripped, one at a time in light-driven reactions (S0 → S4).
- In the recovery step (S4 → S0), O2 is replaced with H2O. 2H2O O M M O2 O O M O O M M M O O M + - O O O M 2H+ 2H + 4e S4 (Mn4O6) S0 (Mn4O4) Adamantane-like Cubane-like - S0, S1, S2 and S3 states take the cubane-like Mn4O4 structure. - S4 states take the adamantane-like structure, i.e., 4O + 2H2O + 4Mn.
- Then, electrons go to substance Z (Tyrosine radical).
O M M O
M O O M
S0 (Mn4O4) Cubane-like
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2. PSII reaction center P680 (Chl a) - P680 = Photon-absorbing center in PSII, which gives the absorption maximum at 680 nm and composed of Chl a’s. - Light causes P680 to be a strong oxidant which accepts electrons from the substance Z via OEC. - Electron flow in PSII: P680 (Chl a) → Chl a → Pheophytin a (Chl a without Mg) → Plastoquinone-Fe (QA) complex → Plastoquinone (QB). - The QB is exchanged with the membrane-bound plastoquinone (Qpool), i.e., the electrons flow into a membrane from PSII. Note: When a plastoquinone (Q) receives two electrons with two protons 2[H•], the plastoquinone becomes QH2 (see page 8). H• = H+ + e3. Electrons to Cyt b6-f complex - 2H+ per electron are transported from stroma to the thylakoid lumen (Total 8 H+). 4. Electrons to plastocyanin (a Cu protein) 5. Electrons to PSI P700 = Photon-absorbing center in PSI, which gives the absorption maximum at 700 nm and composed of Chl a’s. - Photooxidation of P700 yields P700+ (weak oxidant), and subsequently accepts an electron directly from PC. Electrons pass through several carriers (Chl & quinones).
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6. Electrons in PSI flow two possible pathways 1. to NADP+. This is noncyclic pathway. - The electrons are transferred from PSI to the [2Fe-2S] containing soluble ferredoxin (Fd) in stroma, and to the FAD-containing Fd-NADP+ reductase which reduces the NADP+ to NADPH (see Fig. 22-15). 2. back to Cyt b6-f complex. This is a cyclic pathway (see Fig. 22-16). - No NADPH formation, but ATP is produced since H+ is pumped from stroma to thylakoid at Cyt b6-f. Bacteria use this pathway, thus do not produce NADPH. PSI and PSII occupy different parts of the thylakoid membrane - There are three characteristic features: 1. PSI occurs mainly in the unstacked stroma lamella. 2. PSII is located almost exclusively between the closely stacked grana. 3. Cyt b6-f is uniformly distributed.
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This distribution allows chloroplasts to respond to different types of light (full and shady sun).
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Full sun --- more shorter λ light (higher energy) - PSII (P680) absorbs light more than PSI (P700). - Thus PSI cannot handle electrons from PSII. - Therefore, plastoquinones are in a reduced state (PQH2). - The reduced PQH2 activates a protein kinase (PK) which phosphorylates the specific threonine (Thr) of light-harvesting complex (LHC). - The phosphorylated P-LHC migrates to unstacked region of thylakoid membrane where it binds to PSI and funnels the incident light to PSI. - Thus, PSI is activated in order to continue synthesis of NADPH and ATP.
PK (inactive) PQH2 → LHC PK (active) → bind P-LHC ⎯⎯→ PSI (more active)
Shady sun --- more longer λ light (lower energy) - PSI takes electrons faster than PSII produces them since lower energy light reaches less to PSII. - Thus, plastoquinones are mostly in oxidized forms (PQ). - Therefore, LHCs are dephosphorylated, and migrate to PSII. - Thus, the incident light is funneled to PSIIs, and thus PSIIs are activated.
hν
PQH2 protein kinase (PK) dephosphorylation
hν
LHC hν
P
LHC hν
PSII
PQ
PSI
Shady sun
The other reason is that PSII does not act as LHC for PSI.
Full sun
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D. Photophosphorylation (ATP synthesis) - In mitochondria, ΔpH and ΔΨ are used to produce ATP. - The free energy across the membrane is: ΔG = -2.3RT × ΔpH + ZAFΔΨ - In chloroplast, only ΔpH is used (ΔpH =3.5 pH unit). - The free energy across the membrane is: ΔG = -2.3RT × ΔpH - Stroma is basic (low [H+] or high pH) and thylakoid is acidic (high [H+] or low pH). - No electropotential (ΔΨ = 0) across the Chl membrane because the Chl membrane is permeable to Mg2+ (out) and Cl- (in). - When H+’s are transported from stroma to thylakoid lumen, Mg2+’s are oppositely transported. Thus, maintains neutral.
H+ ⎯⎯→ (stroma side) ←⎯⎯ Mg2+ (thylakoid side) Cl ⎯⎯→
ATP productions in noncyclic and cyclic pathways are slightly different - Noncyclic pathway: - 12H+ (4 from 2H2O and 8 from 4PQH2 in Cyt b6-f) are transported by one O2 production, 12H+/O2 or 12H+/(8 absorbed photons). - 3H+ produce one ATP (~ 1 ATP/ 3H+), thus, 4ATP/O2 or 0.5 ATP/ absorbed photon. - Each NADPH produced has the free energy to produce 3ATP. - Thus if the NADPH oxidation is taken into account, 10ATP can be produced by 8 absorbed photons (4ATP by proton gradient and 6ATP by 2NADPH oxidation) or 1.25 ATP per absorbed photon.
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Cyclic pathway: - ~4 additional protons are transported to the thylakoid lumen by the electron from PSI to Cyt b6-f complex. The total H+ transported to thylakoid is 16 (= 12 + 4). - Or, 16H+/8 absorbed photons. - Thus, 16/(8 × 3) = 2/3 ATP per absorbed photon. No. H+ 12 16 No. ATP 4 5 and 1/3 No. NADPH 2 0 ATP/e0.5 (1.25) 2/3 No. e8 8
Noncyclic Cyclic
Note: one photon (hν) = one electron (e-)
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Cartoon representation of light reactions - Let us imagine that PSII and PSI are quite similar to a “windmill”. 1. Four photons hit the propeller of “electron pump” in PSII and rotate it several times. 2. The electron pump in PSII sucks 4 electrons from 2H2O molecules so that the water molecules become O2 and 4H+. 3. The sucked electrons are sent to the Cyt b6-f motor which rotates the “proton pump”. 4. The proton pump transports 8H+ from stroma to thylakoid. 5. The electron voltage is reduced since it is used to rotate the motor of proton pump. 6. Four photons hit the propeller of “electron pump” in PSI and rotate it several times. 7. The electron pump in PSI sucks up electrons from the low voltage to the high voltage. 8a. The activated 4 electrons are used to reduce 2NADP+ to NADPH. 8b. In the cyclic pathway, the activated 4 electrons are sent to the Cyt b6-f motor which rotates the “proton pump” and transports 4H+ from stroma to thylakoid. 9. 4H+ in (2), 8H+ in (4) and 4H+ in (8b if the cyclic pathway) are moved into the proton reservoir of ATP synthesizer. 10. 12H+ or 16H+ (if the cyclic pathway) flow the narrow channel into the ATP synthesizer and rotate the synthesizer’s propeller. 11. The ATP synthesizer phosphorylates 4ADP to produce 4ATP. For the cyclic pathway, 5 and 1/3 ATP are produced.
2NADP cyclic pathway 4hν 4hν 8H+
+
2NADPH
Stroma
4e
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4ADP
+ 4ATP 12H
4e- Electron pump (PSII) O2 + 4H
Motor Electron pump (PSI) 8H
+ +
ATP synthesizer
12H+
2H2O
Proton pump (Cyt b6-f)
Thylakoid lumen
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DARK REACTION It takes place in stroma, and no light is required. It is called “Calvin cycle”. It synthesizes CH2O from CO2 and H2O using pentose phosphate enzymes and other enzymes. The most important other enzyme is ribulose-bisphosphate carboxylase (Rubisco) which catalyzes carboxylation to ribulose-1,5-bisphosphate (RuBP). Overall carboxylation reaction is exergonic (ΔG°’ = -35.1 kJ/mol), which is driven by the cleavage of the β-keto acid intermediate to yield an additional resonance-stabilized carboxylate group.
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Evidence for the above mechanism of ribulose carboxylation. 1. H on C3 of RuBP exchanges with solvent (D2O or T2O), indicating that the hydrogen is acidic. 2. C2 and C3 oxygen atoms remain attached to their respective C atoms, which eliminates mechanisms involving a covalent adduct such as Schiff base between RuBP and the lysine (Lys) residue of enzyme. 3. Carboxylated product, α-keto acid (C1-COO-) can be trapped by NaBH4. But no α-keto is detected. 4. Transition analogue, 2carboxyarabinitol-1-phosphate (CA1P) binds tightly to the enzyme and inhibits the enzyme. 16
Transition state analogues
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Calvin cycle
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Products of light reaction
Rubisco
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Major route of Calvin cycle - is to produce the nucleotide-glucose (XDP-G): Ru5P ⎯ATP→ RuBP ⎯CO2 → 2(3PG) ⎯ATP→ 2(BPG) ⎯NADPH → 2(GAP) ⎯ ⎯ ⎯ ⎯⎯ GAP XTP ⎯→ ⎯ ⎯→ ⎯→ ⎯ GAP ⎯ DHAP ⎯⎯ → FBP ⎯ G6P ⎯ G1P ⎯⎯ → XDP - G where XDP = UDP, ADP, GDP or CDP.
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Products from XDP-G are: Starch ←⎯ XDP-G ⎯→ F6P ⎯→ sucrose-P ⎯→ sucrose
- Others are recovery pathways to re-produce Ru5P.
Calvin cycle can be two stage reactions 1. Stage-1: Production of GAP 3Ru5P + 9ATP + 3CO2 + 6NADPH + 6H+ ⎯→ 6GAP + 6NADP+ + 9ADP + 6Pi 2. Stage-2: Recovery 5GAP ⎯→ 3Ru5P + 2Pi One GAP goes product - Overall reaction: ↑ + 3CO2 + 9ATP + 6NADPH + 6H ⎯→ GAP + 6NADP+ + 9ADP + 8Pi (CO2 + 3ATP + 2NADPH + 2H+ ⎯→ 1 GAP + 2NADP+ + 3ADP + 8/3Pi) 3 B. Control of dark reaction - Major site of control is Rubisco. - During the day, light and dark reactions are satisfactory carried out, but at night, plants must use their nutritional reserves to generate their required ATP and NADPH. - Stroma contains enzymes of glycolysis, oxidative phosphorylation and pentose phosphate pathway as well as Calvin cycle enzymes. - Thus, it is possible to just waste ATP and NADPH by futile cycle, i.e., production of carbohydrates using ATP and NADPH by dark reaction, and reproduction of ATP and NADPH using the carbohydrates by glycolysis, citric acid cycle and oxidative phosphorylation & pentose phosphate pathway. - Therefore, the dark reaction must be control by light sensitive manner. Calvin cycle is activated by light 1. Under light, pH in stroma increases (H+’s are pumped from stroma to thylakoid). - The pH of stroma in night and day are 7.0 and 8.0, respectively. - The optimum pH of Rubisco is 8.0, i.e., thus Rubisco is active only daytime. 2. When H+’s are transported to thylakoid, Mg2+’s are transported to stroma (efflux of Mg2+ to stroma) in order to balance the electric charges. Thus, the [Mg2+] in stroma increases. - Rubisco is activated by Mg2+. 3. Rubisco is allosterically activated by NADPH produced by light reaction. - At low [NADPH] (i.e., at night time), Rubisco is less active. 4. The potent inhibitor and transition analog of Rubisco, 2-carboxyarabinitol-1-phosphate (CA1P) is only synthesized in the dark. - Thus Rubisco is inhibited by CA1P in night.
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Second regulatory system (other than Rubisco regulation)---Mainly day time regulation - In recovery stage, two phosphatases are activated by increased pH, Mg2+ and NADPH. 1. Fructose bisphosphatase, FBPase at step-7. 2. Sedoheptulose bisphosphatase, SBPase at step-10. - Actually, FBPase and SBPase are activated by the reduced ferredoxin (Fd) via thioredoxin reductase. - The reduced Fd also inhibits PFK. - Thus, glycolysis is inhibited and gluconeogenesis is stimulated in day time.
Inhibits PFK
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Photorespiration and the C4 cycle - Rubisco can use O2 instead of CO2, since the KM values of [O2] and [CO2] in plants are nearly the same. - When Rubisco uses O2, its process is called “photorespiration”. Photorespiration
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Rubisco ⎯→ RuBP + O2 ⎯⎯⎯⎯ 3PG + 2-phosphoglycolate (C2) The proposed reaction mechanism:
Carboxylation vs. oxygenation - Carboxylation: RuBP + CO2 → 2(3PG) - Oxygenation: RuBP + O2 → 3PG + C2 (2-phosphoglycolate) ⏐ ⏐NADH + ATP ⏐ ↓ CO2
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Photorespiration is taken place in three organelles (Chloroplast, Peroxisome and Mitochondrion) The 2-phosphoglycolate produced by RuBP oxygenation is dephosphorylated, and is moved into a peroxisome (glyoxisome).
At there, glycolate is converted to glycine (Gly) by receiving NH3. The Gly is transported into a mitochondrion. At there, two Gly are condensed into serine (Ser) and CO2. The Ser is re-transported into a peroxisome. At there, the Ser is deaminated and becomes glycerate. The glycerate is phosphorylated by ATP in cytosol, and becomes 3PG. The 3PG is re-transported into chloroplast, and enters the Calvin
cycle.
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The overall photorespiration is: 2RuBP + O2 → 2(3PG) + Ser + CO2 then Ser + 3PG + NADPH + ATP → RuBP + CO2 + NADP+ + ADP The overall processes are: RuBP + O2 + NADPH + ATP → 3PG + 2CO2 + NADP+ + ADP Thus, photorespiration is detrimental wasteful process.
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Some plants have a way to partially overcome O2 reaction - This process is called “C4 cycle”. - Photosynthesis takes place in bundle sheath cells where Calvin cycle occurs. - In C4 plants, bundle sheath cells are covered by mesophyll cells which do not contain Rubisco. Thus, the photorespiration does not occur in mesophyll cells. - In mesophyll cells, - Pyruvate is converted to PEP by consumption of two ATP. - CO2 is fixed on PEP to produce oxaloacetate (C4 molecule). - Reduced oxaloacetate, malate is transported to bundle sheath cells. - In bundle sheath cells, - Then the malate is reoxidized to pyruvate and CO2. - The CO2 is entered into Calvin cycle.
No Rubisco
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C4 plants require five ATP to fix one CO2 in their photosynthesis (C4 cycle + Calvin cycle), where as C3 plants use three ATP to fix one CO2. Note: C3 plants initially fix CO2 in the form of three-carbon acids, such as 3PG. C4 plants initially fix CO2 in the form of four-carbon acids, such as oxaloacetate. C4 plants have advantage in warm climate, since the stomata closed much of time to avoid H2O loss. These plants absorb CO2 at night and store it as malate, and the stored CO2 is used at day time. Mesophyll cells also prevent from O2 to access to the bundle-sheath cells.
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CAM plants (Crassulacean acid metabolism) - C4 plants separate CO2 fixation (Calvin cycle) by space, i.e., mesophyll cells. - CAM plants do not have mesophyll cells, but their stomata are closed during day to prevent loss of H2O. Thus, O2 cannot enter in day time.
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At night (Rubisco activity is low), stomata is opened to absorb H2O and CO2. CO2 is stored in PEP as malate as C4 plants. CO2 + PEP → OAA → Malate (stored) At daytime, CO2 is regenerated from the stored malate, and enters into Calvin cycle to produce CH2O. Calvin cycle ⎯ Malate → CO2 + PEP and CO2 + RuBP ⎯ ⎯⎯⎯⎯ → 3PG Thus, CAM plants separate CO2 fixation (Calvin cycle) in time (day and night).
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Takusagawa’s Note©
Chapter 18: Photosynthesis
1. CHLOROPLASTS - Photosynthesis is carried out at chloroplasts. Structure of chloroplast
Outer membrane Stroma lamellae Inner membrane Thylakoid
Chloroplast
Granum Dark reaction Stroma Light reaction
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Composition of innermembrane and granum membrane are unusual. - Phospholipid (negatively charged) ~10% - Neutral lipid (galactose) ~80%
Photosynthesis occurs in two distinct phases: 1. Light reactions --- Generates NADPH & ATP by using light energy at thylakoid membrane (prokaryotes: inner membrane). 2. Dark reactions --- Synthesize carbohydrates from CO2 + H2O by using NADPH & ATP at stroma.
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LIGHT REACTION Initially believed: CO2 + H2O ⎯light → CH2O + O2 ⎯ van Niel showed: CO2 + 2H2S ⎯light → CH2O + 2S + H2O ⎯ In general: CO2 + 2H2A ⎯light → CH2O + 2A + H2O ⎯
This suggested that photosynthesis is two step reactions: 1. Light energy oxidizes H2A (light reaction): 2H2A ⎯light → 2A + 4[H] ⎯ 2. The reducing agent [H] reduces CO2 (dark reaction): 4[H] + CO2 ⎯→ CH2O + H2O
A. Absorption of light - The principal photoreceptor is chlorophyll (Chl) derived from protoporphyrin IX. - Major differences between chlorophyll and hemes are: - Chl has an extra ring (V) (cyclopentanone). - Ring IV of Chl is more reduced. - …show more content…
Mg is present instead of Fe. - Side chains are different --- some Chl have a long chain at R4 site.
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Chapter 18 Eukaryotes have: - Chlorophyll a (Chl a) - Chlorophyll b (Chl b) Prokaryotes have: - Bacteriochlorophyll a (BChl a) - Bacteriochlorophyll b (BChl b)
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Takusagawa’s Note©
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- Note: No absorption of Chl a and b in 480 - 620 nm region. But other pigments, carotenoids (such as β-carotene), phycoerythrin and phycocyanin absorb the light between 480 and 620.
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Chapter 18 Photo-energy Photo-energy E = hν = hc
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Takusagawa’s Note©
light (2.998 × 108 m/s), and λ = wavelength of light. - One mole of red light (6.02 × 1023 photons = 1 einstein) with λ= 700 nm has 171 kJ/einstein: Nhc (6 × 10 23 )(6.6 × 10 −34 J ⋅ s −1 )(3 × 10 8 m ⋅ s −1 ) E= = = 171kJ / einstein . λ 700 × 10 −9 m - In general, when molecules absorb photon energy, they move from ground state to various excited states. - There are four pathways that an excited state returns to lower energy state. 1. Losing energy as heat (kinetic). 2. Losing energy as light (fluorescence) that normally shifts to longer wavelength. 3*. Losing energy as energy transfer to another molecule. ⎯ MA + MB ⎯light → M*A + MB ⎯→ MA + M*B * 4 . Losing energy by losing electron (photooxidation) ⎯ Chl ⎯light → Chl+ + e* 3 and 4 are important for photosynthesis.
λ
, where h = Planck’s constant (6.626 × 10-34 J/s), c = speed of
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Takusagawa’s Note©
Light absorbed by antenna chlorophylls and accessory pigments is transferred to photosynthetic reaction centers - Light is absorbed in “light-harvesting complexes” (LHCs) located on the membrane of thylakoid in chloroplast. - LHC is membrane bound proteins containing Chl molecules and other pigments.
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Light-harvesting complex in pea chloroplast Most Chl molecules and other pigment molecules absorb light and the light energy (exciton) is randomly transferred to the neighboring Chl or other pigment until it is trapped by a “photosynthetic reaction center”. Thus, the function of most Chl’s is to gather light, i.e., they act as light-harvesting antennas. Photosynthetic reaction centers contain the same Chl molecules as antennas have. However, Chl molecules in photosynthetic reaction centers have slightly lower excited state energies because their different environments. Thus, excitons are trapped or funneled into reaction centers. A photosynthetic reaction center is surrounded by ~300 photosynthetic antenna complexes. Photosynthetic reaction center
Schemetic diagram of light harvesting complex
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Chloroplast
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Takusagawa’s Note©
C. Plant Photosynthesis is two-center electron transport - Bacteria photosynthesis is one center process, i.e., one light absorbing event, whereas - Photosynthesis in plants and algae is two center process, i.e., two light absorbing event. light (4 photons) 1. 2H2O ⎯⎯⎯⎯⎯⎯⎯ ⎯→ O2 + 4e- + 4H+ light (4 photons) ⎯ 2. 2NADP+ + 4e- + 2H+ ⎯ ⎯⎯⎯⎯⎯⎯ → 2NADPH Thus, light is used to reduce NADP+ with H2O. 2NADP+ + 2H2O ⎯→ 2NADPH + O2 + 2H+
ΔE°′ = -0.815 V ΔE°′ = -0.320 V ΔE°’ = -1.135 V
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ΔG°’ = -nFΔE°’ = 438 kJ/mol, where n = 4 (since 4 electrons are involved) and F=96.5 kJ/V.
As explained in previously, photo-energy of 700 nm light per one mole of photon (einstein) is: E = 171 kJ/einstein. In each event, four electrons are involved. Thus the total electrons are eight (Remember: two-light absorbing events). One electron activation requires one photon. Thus eight photons are necessary to activate eight electrons. The input light energy is: E = 8 × 171 = 1368 kJ/mol, which is much larger than ΔG°’(438 kJ/mol) of 2 mol NADPH and 1 mol O2 productions.
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-
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ΔG°’ can be calculated from the pair of half-cell reactions listed in Table 15-4. NADP+ + H+ + 2e- → NADPH E°’ = -0.320 V + H2O → ½O2 + 2H + 2e E°’ = -0.815 V Thus, 2 mol NADPH and 1 mol O2 production require 2 × (-0.320 + -0.815) = -2.27 V. Since ΔG°’ = -nFΔE°’, ΔG°’ = -2 × 96.5 × -2.27 = 438.11 kJ/mol
Photosynthetic O2 production requires two-sequential photosystems - These events are occurred in two different photosynthetic reaction centers that are connected essentially in series. 1. Photosystem II (PSII) --- generates a weak reductant and strong oxidant to oxidize H2O. light (4 photons) 2H2O ⎯⎯⎯⎯⎯⎯⎯ → O2 + 4e- + 4H+ ⎯
eChl light Chl* Chl+ e- (from H2O)
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8
Takusagawa’s Note©
2. Photosystem I (PSI) --- generates a strong reductant to reduce NADP+ and weak oxidant. light (4 photons) 2NADP+ + 4e- + 2H+ ⎯ ⎯⎯⎯⎯⎯⎯ → 2NADPH ⎯
Chl
light
eChl* Chl-
(to NADP+) e
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Herbicide dichlorophenyl-dimethylurea (DCMU) blocks photosynthetic electron transport from PSII to cytochrome b6-f.
8
Chapter 18
9
Takusagawa’s Note©
Three complexes in membrane - are involved in the electron transport from (2H2O → O2 + 4H+ + 4e-) event to (2NADP+ + 2H+ + 4e- → 2NADPH) event. Those are: 3) PSI 1) PSII 2) Cyt b6-f complex - The electrons are carried by the mobile electron carriers between the complexes. Plastoquinol (PQ) --- between PSII and Cyt b6-f complex. Plastocyanin (PC) ---between Cyt b6-f complex and PSI. - Electron flow: PSII → (PQ) → Cyt b6-f complex → (PC) → PSI
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Plastoquinol (PQ) has two oxidation states, PQ (oxidized state) and PQH2 (reduced state). Plastocyanin (PC) is a small Cu-containing protein.
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10
Takusagawa’s Note©
Details of electron transport 1. Oxidation of H2O by “O2-evolving complex” (OEC) in PSII. - OEC contains 4Mn cluster that binds 2H2O. - Oxidation of H2O occurs 5 steps, i.e., S0, S1, S2, S3, S4. Four electrons are stripped, one at a time in light-driven reactions (S0 → S4).
- In the recovery step (S4 → S0), O2 is replaced with H2O. 2H2O O M M O2 O O M O O M M M O O M + - O O O M 2H+ 2H + 4e S4 (Mn4O6) S0 (Mn4O4) Adamantane-like Cubane-like - S0, S1, S2 and S3 states take the cubane-like Mn4O4 structure. - S4 states take the adamantane-like structure, i.e., 4O + 2H2O + 4Mn.
- Then, electrons go to substance Z (Tyrosine radical).
O M M O
M O O M
S0 (Mn4O4) Cubane-like
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11
Takusagawa’s Note©
2. PSII reaction center P680 (Chl a) - P680 = Photon-absorbing center in PSII, which gives the absorption maximum at 680 nm and composed of Chl a’s. - Light causes P680 to be a strong oxidant which accepts electrons from the substance Z via OEC. - Electron flow in PSII: P680 (Chl a) → Chl a → Pheophytin a (Chl a without Mg) → Plastoquinone-Fe (QA) complex → Plastoquinone (QB). - The QB is exchanged with the membrane-bound plastoquinone (Qpool), i.e., the electrons flow into a membrane from PSII. Note: When a plastoquinone (Q) receives two electrons with two protons 2[H•], the plastoquinone becomes QH2 (see page 8). H• = H+ + e3. Electrons to Cyt b6-f complex - 2H+ per electron are transported from stroma to the thylakoid lumen (Total 8 H+). 4. Electrons to plastocyanin (a Cu protein) 5. Electrons to PSI P700 = Photon-absorbing center in PSI, which gives the absorption maximum at 700 nm and composed of Chl a’s. - Photooxidation of P700 yields P700+ (weak oxidant), and subsequently accepts an electron directly from PC. Electrons pass through several carriers (Chl & quinones).
11
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12
Takusagawa’s Note©
6. Electrons in PSI flow two possible pathways 1. to NADP+. This is noncyclic pathway. - The electrons are transferred from PSI to the [2Fe-2S] containing soluble ferredoxin (Fd) in stroma, and to the FAD-containing Fd-NADP+ reductase which reduces the NADP+ to NADPH (see Fig. 22-15). 2. back to Cyt b6-f complex. This is a cyclic pathway (see Fig. 22-16). - No NADPH formation, but ATP is produced since H+ is pumped from stroma to thylakoid at Cyt b6-f. Bacteria use this pathway, thus do not produce NADPH. PSI and PSII occupy different parts of the thylakoid membrane - There are three characteristic features: 1. PSI occurs mainly in the unstacked stroma lamella. 2. PSII is located almost exclusively between the closely stacked grana. 3. Cyt b6-f is uniformly distributed.
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This distribution allows chloroplasts to respond to different types of light (full and shady sun).
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Takusagawa’s Note©
Full sun --- more shorter λ light (higher energy) - PSII (P680) absorbs light more than PSI (P700). - Thus PSI cannot handle electrons from PSII. - Therefore, plastoquinones are in a reduced state (PQH2). - The reduced PQH2 activates a protein kinase (PK) which phosphorylates the specific threonine (Thr) of light-harvesting complex (LHC). - The phosphorylated P-LHC migrates to unstacked region of thylakoid membrane where it binds to PSI and funnels the incident light to PSI. - Thus, PSI is activated in order to continue synthesis of NADPH and ATP.
PK (inactive) PQH2 → LHC PK (active) → bind P-LHC ⎯⎯→ PSI (more active)
Shady sun --- more longer λ light (lower energy) - PSI takes electrons faster than PSII produces them since lower energy light reaches less to PSII. - Thus, plastoquinones are mostly in oxidized forms (PQ). - Therefore, LHCs are dephosphorylated, and migrate to PSII. - Thus, the incident light is funneled to PSIIs, and thus PSIIs are activated.
hν
PQH2 protein kinase (PK) dephosphorylation
hν
LHC hν
P
LHC hν
PSII
PQ
PSI
Shady sun
The other reason is that PSII does not act as LHC for PSI.
Full sun
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14
Takusagawa’s Note©
D. Photophosphorylation (ATP synthesis) - In mitochondria, ΔpH and ΔΨ are used to produce ATP. - The free energy across the membrane is: ΔG = -2.3RT × ΔpH + ZAFΔΨ - In chloroplast, only ΔpH is used (ΔpH =3.5 pH unit). - The free energy across the membrane is: ΔG = -2.3RT × ΔpH - Stroma is basic (low [H+] or high pH) and thylakoid is acidic (high [H+] or low pH). - No electropotential (ΔΨ = 0) across the Chl membrane because the Chl membrane is permeable to Mg2+ (out) and Cl- (in). - When H+’s are transported from stroma to thylakoid lumen, Mg2+’s are oppositely transported. Thus, maintains neutral.
H+ ⎯⎯→ (stroma side) ←⎯⎯ Mg2+ (thylakoid side) Cl ⎯⎯→
ATP productions in noncyclic and cyclic pathways are slightly different - Noncyclic pathway: - 12H+ (4 from 2H2O and 8 from 4PQH2 in Cyt b6-f) are transported by one O2 production, 12H+/O2 or 12H+/(8 absorbed photons). - 3H+ produce one ATP (~ 1 ATP/ 3H+), thus, 4ATP/O2 or 0.5 ATP/ absorbed photon. - Each NADPH produced has the free energy to produce 3ATP. - Thus if the NADPH oxidation is taken into account, 10ATP can be produced by 8 absorbed photons (4ATP by proton gradient and 6ATP by 2NADPH oxidation) or 1.25 ATP per absorbed photon.
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Cyclic pathway: - ~4 additional protons are transported to the thylakoid lumen by the electron from PSI to Cyt b6-f complex. The total H+ transported to thylakoid is 16 (= 12 + 4). - Or, 16H+/8 absorbed photons. - Thus, 16/(8 × 3) = 2/3 ATP per absorbed photon. No. H+ 12 16 No. ATP 4 5 and 1/3 No. NADPH 2 0 ATP/e0.5 (1.25) 2/3 No. e8 8
Noncyclic Cyclic
Note: one photon (hν) = one electron (e-)
14
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15
Takusagawa’s Note©
Cartoon representation of light reactions - Let us imagine that PSII and PSI are quite similar to a “windmill”. 1. Four photons hit the propeller of “electron pump” in PSII and rotate it several times. 2. The electron pump in PSII sucks 4 electrons from 2H2O molecules so that the water molecules become O2 and 4H+. 3. The sucked electrons are sent to the Cyt b6-f motor which rotates the “proton pump”. 4. The proton pump transports 8H+ from stroma to thylakoid. 5. The electron voltage is reduced since it is used to rotate the motor of proton pump. 6. Four photons hit the propeller of “electron pump” in PSI and rotate it several times. 7. The electron pump in PSI sucks up electrons from the low voltage to the high voltage. 8a. The activated 4 electrons are used to reduce 2NADP+ to NADPH. 8b. In the cyclic pathway, the activated 4 electrons are sent to the Cyt b6-f motor which rotates the “proton pump” and transports 4H+ from stroma to thylakoid. 9. 4H+ in (2), 8H+ in (4) and 4H+ in (8b if the cyclic pathway) are moved into the proton reservoir of ATP synthesizer. 10. 12H+ or 16H+ (if the cyclic pathway) flow the narrow channel into the ATP synthesizer and rotate the synthesizer’s propeller. 11. The ATP synthesizer phosphorylates 4ADP to produce 4ATP. For the cyclic pathway, 5 and 1/3 ATP are produced.
2NADP cyclic pathway 4hν 4hν 8H+
+
2NADPH
Stroma
4e
-
4ADP
+ 4ATP 12H
4e- Electron pump (PSII) O2 + 4H
Motor Electron pump (PSI) 8H
+ +
ATP synthesizer
12H+
2H2O
Proton pump (Cyt b6-f)
Thylakoid lumen
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3. -
16
Takusagawa’s Note©
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DARK REACTION It takes place in stroma, and no light is required. It is called “Calvin cycle”. It synthesizes CH2O from CO2 and H2O using pentose phosphate enzymes and other enzymes. The most important other enzyme is ribulose-bisphosphate carboxylase (Rubisco) which catalyzes carboxylation to ribulose-1,5-bisphosphate (RuBP). Overall carboxylation reaction is exergonic (ΔG°’ = -35.1 kJ/mol), which is driven by the cleavage of the β-keto acid intermediate to yield an additional resonance-stabilized carboxylate group.
-
Evidence for the above mechanism of ribulose carboxylation. 1. H on C3 of RuBP exchanges with solvent (D2O or T2O), indicating that the hydrogen is acidic. 2. C2 and C3 oxygen atoms remain attached to their respective C atoms, which eliminates mechanisms involving a covalent adduct such as Schiff base between RuBP and the lysine (Lys) residue of enzyme. 3. Carboxylated product, α-keto acid (C1-COO-) can be trapped by NaBH4. But no α-keto is detected. 4. Transition analogue, 2carboxyarabinitol-1-phosphate (CA1P) binds tightly to the enzyme and inhibits the enzyme. 16
Transition state analogues
Chapter 18
17
Calvin cycle
Takusagawa’s Note©
Products of light reaction
Rubisco
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Takusagawa’s Note©
Major route of Calvin cycle - is to produce the nucleotide-glucose (XDP-G): Ru5P ⎯ATP→ RuBP ⎯CO2 → 2(3PG) ⎯ATP→ 2(BPG) ⎯NADPH → 2(GAP) ⎯ ⎯ ⎯ ⎯⎯ GAP XTP ⎯→ ⎯ ⎯→ ⎯→ ⎯ GAP ⎯ DHAP ⎯⎯ → FBP ⎯ G6P ⎯ G1P ⎯⎯ → XDP - G where XDP = UDP, ADP, GDP or CDP.
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Products from XDP-G are: Starch ←⎯ XDP-G ⎯→ F6P ⎯→ sucrose-P ⎯→ sucrose
- Others are recovery pathways to re-produce Ru5P.
Calvin cycle can be two stage reactions 1. Stage-1: Production of GAP 3Ru5P + 9ATP + 3CO2 + 6NADPH + 6H+ ⎯→ 6GAP + 6NADP+ + 9ADP + 6Pi 2. Stage-2: Recovery 5GAP ⎯→ 3Ru5P + 2Pi One GAP goes product - Overall reaction: ↑ + 3CO2 + 9ATP + 6NADPH + 6H ⎯→ GAP + 6NADP+ + 9ADP + 8Pi (CO2 + 3ATP + 2NADPH + 2H+ ⎯→ 1 GAP + 2NADP+ + 3ADP + 8/3Pi) 3 B. Control of dark reaction - Major site of control is Rubisco. - During the day, light and dark reactions are satisfactory carried out, but at night, plants must use their nutritional reserves to generate their required ATP and NADPH. - Stroma contains enzymes of glycolysis, oxidative phosphorylation and pentose phosphate pathway as well as Calvin cycle enzymes. - Thus, it is possible to just waste ATP and NADPH by futile cycle, i.e., production of carbohydrates using ATP and NADPH by dark reaction, and reproduction of ATP and NADPH using the carbohydrates by glycolysis, citric acid cycle and oxidative phosphorylation & pentose phosphate pathway. - Therefore, the dark reaction must be control by light sensitive manner. Calvin cycle is activated by light 1. Under light, pH in stroma increases (H+’s are pumped from stroma to thylakoid). - The pH of stroma in night and day are 7.0 and 8.0, respectively. - The optimum pH of Rubisco is 8.0, i.e., thus Rubisco is active only daytime. 2. When H+’s are transported to thylakoid, Mg2+’s are transported to stroma (efflux of Mg2+ to stroma) in order to balance the electric charges. Thus, the [Mg2+] in stroma increases. - Rubisco is activated by Mg2+. 3. Rubisco is allosterically activated by NADPH produced by light reaction. - At low [NADPH] (i.e., at night time), Rubisco is less active. 4. The potent inhibitor and transition analog of Rubisco, 2-carboxyarabinitol-1-phosphate (CA1P) is only synthesized in the dark. - Thus Rubisco is inhibited by CA1P in night.
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Takusagawa’s Note©
Second regulatory system (other than Rubisco regulation)---Mainly day time regulation - In recovery stage, two phosphatases are activated by increased pH, Mg2+ and NADPH. 1. Fructose bisphosphatase, FBPase at step-7. 2. Sedoheptulose bisphosphatase, SBPase at step-10. - Actually, FBPase and SBPase are activated by the reduced ferredoxin (Fd) via thioredoxin reductase. - The reduced Fd also inhibits PFK. - Thus, glycolysis is inhibited and gluconeogenesis is stimulated in day time.
Inhibits PFK
19
Chapter 18
20
Takusagawa’s Note©
Photorespiration and the C4 cycle - Rubisco can use O2 instead of CO2, since the KM values of [O2] and [CO2] in plants are nearly the same. - When Rubisco uses O2, its process is called “photorespiration”. Photorespiration
-
Rubisco ⎯→ RuBP + O2 ⎯⎯⎯⎯ 3PG + 2-phosphoglycolate (C2) The proposed reaction mechanism:
Carboxylation vs. oxygenation - Carboxylation: RuBP + CO2 → 2(3PG) - Oxygenation: RuBP + O2 → 3PG + C2 (2-phosphoglycolate) ⏐ ⏐NADH + ATP ⏐ ↓ CO2
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Chapter 18
21
Takusagawa’s Note©
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Photorespiration is taken place in three organelles (Chloroplast, Peroxisome and Mitochondrion) The 2-phosphoglycolate produced by RuBP oxygenation is dephosphorylated, and is moved into a peroxisome (glyoxisome).
At there, glycolate is converted to glycine (Gly) by receiving NH3. The Gly is transported into a mitochondrion. At there, two Gly are condensed into serine (Ser) and CO2. The Ser is re-transported into a peroxisome. At there, the Ser is deaminated and becomes glycerate. The glycerate is phosphorylated by ATP in cytosol, and becomes 3PG. The 3PG is re-transported into chloroplast, and enters the Calvin
cycle.
-
The overall photorespiration is: 2RuBP + O2 → 2(3PG) + Ser + CO2 then Ser + 3PG + NADPH + ATP → RuBP + CO2 + NADP+ + ADP The overall processes are: RuBP + O2 + NADPH + ATP → 3PG + 2CO2 + NADP+ + ADP Thus, photorespiration is detrimental wasteful process.
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Chapter 18
22
Takusagawa’s Note©
Some plants have a way to partially overcome O2 reaction - This process is called “C4 cycle”. - Photosynthesis takes place in bundle sheath cells where Calvin cycle occurs. - In C4 plants, bundle sheath cells are covered by mesophyll cells which do not contain Rubisco. Thus, the photorespiration does not occur in mesophyll cells. - In mesophyll cells, - Pyruvate is converted to PEP by consumption of two ATP. - CO2 is fixed on PEP to produce oxaloacetate (C4 molecule). - Reduced oxaloacetate, malate is transported to bundle sheath cells. - In bundle sheath cells, - Then the malate is reoxidized to pyruvate and CO2. - The CO2 is entered into Calvin cycle.
No Rubisco
-
C4 plants require five ATP to fix one CO2 in their photosynthesis (C4 cycle + Calvin cycle), where as C3 plants use three ATP to fix one CO2. Note: C3 plants initially fix CO2 in the form of three-carbon acids, such as 3PG. C4 plants initially fix CO2 in the form of four-carbon acids, such as oxaloacetate. C4 plants have advantage in warm climate, since the stomata closed much of time to avoid H2O loss. These plants absorb CO2 at night and store it as malate, and the stored CO2 is used at day time. Mesophyll cells also prevent from O2 to access to the bundle-sheath cells.
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Takusagawa’s Note©
CAM plants (Crassulacean acid metabolism) - C4 plants separate CO2 fixation (Calvin cycle) by space, i.e., mesophyll cells. - CAM plants do not have mesophyll cells, but their stomata are closed during day to prevent loss of H2O. Thus, O2 cannot enter in day time.
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At night (Rubisco activity is low), stomata is opened to absorb H2O and CO2. CO2 is stored in PEP as malate as C4 plants. CO2 + PEP → OAA → Malate (stored) At daytime, CO2 is regenerated from the stored malate, and enters into Calvin cycle to produce CH2O. Calvin cycle ⎯ Malate → CO2 + PEP and CO2 + RuBP ⎯ ⎯⎯⎯⎯ → 3PG Thus, CAM plants separate CO2 fixation (Calvin cycle) in time (day and night).
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23