Energy conservation in photosynthesis: Harvesting Sunlight

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4 Energy conservation in photosynthesis
Harvesting Sunlight
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The primary function of leaves is photosynthesis.
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Focus of this chapter (1)

• The structure of higher plant leaves with respect
  to the interception of light
• Photosynthesis as the reduction of carbon dioxide
  to carbohydrate
• The photosynthetic electron transport chain, its
  organization in the thylakoid membrane, and its
  role in generating reducing potential and ATP
• Problems encounters by chloroplasts when they are
  subjected to varying amount of light

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Focus of this chapter (2)

• The dynamic nature of the thylakoid membrane,
  showing how changes in the organization of
  light-harvesting apparatus influence the
  absorption and distribution of light energy
• The role of carotenoids as accessory pigments and
  in photoprotection of chlorophyll and
• The use of herbicides that specifically interact
  with photosynthetic electron transport

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The structure of the leaf

• The architecture of a typical higher plant leaf
  is particularly well suited to absorb light.
• The photosynthetic tissues (mesophyll) are
  located between the two epidermal layers.
• Dicotyledonous leaf is structurally different
  from monocotyledonous leaf.

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The structure of dicotyledonous leaf

• One-to-three layers of palisade mesophyll cells
  forms the upper photosynthetic tissue.
• Below is the spongy mesophyll cells.

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The structure of dicotyledonous leaf

• Palisade mesophyll cells are elongated,
  cylindrical cells with the long axis
  perpendicular to the surface of the leaf.
• Spongy mesophyll cells are irregular with lots of
  air spaces between the cells.

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The structure of monocotyledonous leaf

• Monocotyledonous leaf lack the distinction
  between palisade and spongy mesophyll cells.

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Comparison between mesophyll cells

• Palisade mesophyll generally have larger numbers
  of chloroplasts than spongy mesophyll.

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Sieve effect

• When light passes through the first layer of
  cells (palisade mesophyll cells) without being
  absorbed, we call this sieve effect.
• The sieve effect is due to the fact that
  chlorophyll is not uniformly distributed
  throughout cells but instead is confined to the
  chloroplasts.

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Sieve effect

• To reduce sieve effect, plant develops multiple
  layers of photosynthetic cells.
• The reflection, refraction, and scattering of
  light inside leaf may also reduce sieve effect.

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Photosynthesis

• Photosynthesis can be viewed as a photochemical
  reduction of CO2.
• In the 1920s, C.B. van Niel discovered the O2
  produced from photosynthesis is from water.
• In 1939 Robert Hill found light reaction still
  can happen in isolated chloroplast when no CO2 is
  consumed and no carbohydrate was produced.
• In the early 1940s S. Ruben and M. Kamen showed
  O2 produced from photosynthesis is from water by
  using O18 labeled water.

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Photosynthetic electron transport

• The principle function of the light-dependent
  reactions of photosynthesis is therefore to
  generate the NADPH and ATP required for carbon
  reduction.

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Photosynthetic electron transport

• The effect of photosynthetic electron transport
  chain is to extract low-energy electrons from
  water and raise the energy level of those
  electrons to produce a strong reductant NADPH.
• The energy plant used to raise the energy level
  of those electrons is the light energy trapped by
  chlorophyll.

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Photosynthetic electron-transport chain

• Two large, multimolecular complexes, photosystem
  I (PSI) and photosystem II (PSII), linked with a
  third multiprotein aggregate called the
  cytochrome complex, form the photosynthetic
  electron-transport chain.

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Photosystems

• Photosystems contain several different proteins
  together with a collection of chlorophyll and
  carotenoid molecules that absorb photons.
• Most of the chlorophyll in the photosystem
  functions as antenna chlorophyll.

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Photosystems

• The antenna chlorophyll absorb light but do not
  participate in photochemical reactions. It pass
  its energy to the next chlorophyll by either
  inductive resonance or radiationless energy
  transfer.

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Reaction center of photosystem

• For PSII, each reaction center consisted of two
  chlorophyll a called reaction center chlorophyll.
• Reaction center chlorophyll is the lowest-energy
  absorbing chlorophyll in the PSII complex (energy
  sink).

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Energy transfer efficiency of Photosystem

• The design of photosystems ensure efficient
  energy transfer. Only about 10 of the energy is
  lost during the whole transfer process (from
  antenna to reaction center chlorophyll).

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Why photosystems?

• The principle advantage of associating a single
  reaction center with a large number of antenna
  chlorophyll molecules is to increase efficiency
  in the collection and utilization of light energy.

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Why photosystems?

• Even in bright sunlight, an individual
  chlorophyll will only be struck not more than a
  few times per second. However, energy transfer
  only takes ms. So it is more economical not to
  make every chlorophyll into reaction center.

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Light-harvesting complexes (LHC) are closely
associated with photosystems
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Light-harvesting complexes (LHC)

• Light harvesting complex (also consisted of
  chlorophyll and proteins) serves as extended
  antenna systems for harvesting additional light
  energy.
• In chloroplast, there are two LHCs. The one
  associated with PSI is named LHCI and the one
  associated with PSII is named LHCII, accordingly.

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Light-harvesting complexes (LHC)

• All the chlorophyll b are contained in LHCs. Most
  of the chloroplast pigments (70) are in LHCs.
• LHCI has a chlorophyll a/b ratio about 4 and it
  is tightly bound to PSI.
• LHCII has a chlorophyll a/b ratio about 1.2.
  Besides owning most of the chloroplast
  chlorophyll (5060), LHCII also contains most of
  the chlorophyll b and xanthophyll.

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Photosynthetic electron transport chain
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PSII ? pheophytin

• P680 is located at the lumenal side of reaction
  center.
• When excited, the excited P680 (P680) is rapidly
  (10-12s) photooxidized as it passes an electron
  to pheophytin (primary electron acceptor).

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pheophytin

• Pheophytin is a form of chlorophyll a with the
  Mg2 replaced by two hydrogens.
• The photo-oxidation of P680 is then followed by
  charge separation (P680Pheo-).

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phytyl
Pheophytin
Pheophytin a R1 -CH3 R2 phytyl Pheophytin b
R1 -CHO R2 phytyl
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P680 ? pheophytin

• Noted the direction of electron movement in PSII.
  P680 is located at the lumen side of PSII, then
  the electron is transferred to pheophytin, which
  is more towards the stromal side, so electron
  will not recombine with P680.

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Pheophytin ? QA ? PQ

• Reaction proteins D1 and D2 orient specific redox
  carriers of the PSII reaction center so the
  probability of charge recombination is further
  reduced.

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Pheophytin ? QA ? PQ

• D2 contains QA (quinone electron acceptor) which
  will accept electrons from pheophytin within
  picoseconds.
• Then electron from QA will be passed to
  plastoquinone (PQ), a quinone bound transiently
  to the binding site on D1 protein (QB).

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Plastoquinone (PQ)

• The reduction of plastoquinone (PQ) to
  plastoquinol (PQH2) lowering the affinity of this
  molecule for the binding site.
• After plastoquinol is released from the reaction
  center, another molecule of PQ will occupy the
  empty space.

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Oxygen-evolving complex (OEC)

• P680 got its electron directly from a cluster of
  four Mn2 associated with a small complex of
  proteins.
• OEC is located on the lumen side of the thylakoid
  membrane and bound to the D1 and D2 proteins of
  PSII reaction center.

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Oxygen-evolving complex (OEC)

• The OEC proteins functions to stabilize the Mn2
  cluster.
• Chloride ion (Cl-) is also required for the water
  splitting function.

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Oxygen-evolving complex (OEC)

• To generate one molecule of O2, four electrons
  must be withdrawn from two molecule of H2O. This
  suggest that OEC should be able to store charges
  (and experiment results agree with this).

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PQ ? cyt b6f complex

• After plastoquinol is released from PSII, it
  diffuses through the membrane until reaches
  cytochrome b6f complex.
• Because plastoquinol has to reach cyt b6f by
  diffusion, this is the slowest step in
  photosynthetic electron transport (milliseconds).

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Cytb6f complex

• Electron is then transferred from plastoquinol to
  Rieske iron-sulfur (FeS) protein ? cytochrome f
  (all on the lumenal side).
• Then electrons are picked up by plastocyanin (PC).

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Plastocyanin (PC)

• Plastocyanin is a small peripheral protein that
  is able to diffuse freely along the lumenal
  surface of the thylakoid membrane.

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PC ? PSI

• PC is then transfer electron to PSI.
• The reaction center chlorophyll (P700) first
  become P700, then photooxidized to P700 and
  give its electron to a molecule of chlorophyll a.

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Photosystem I

• The electron is then passed to a quinone
  (phylloquinone).
• Electron transfer then proceeds through a series
  of Fe-S centers and ultimately to the soluble
  iron-sulfur protein, ferredoxin (Fdx).

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Ferredoxin ? NADP

• Ferredoxin-NADP reductase (Fd-NADP reductase)
  then uses ferredoxin to reduce NADP.

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Although PSI do accept electrons from
plastocyanin, PSI
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can also be activated by light.

• When PSI is directly activated by light, the
  electron it lost is satisfied by withdrawing an
  electron from reduced PC.

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Photosynthetic efficiency

• The efficiency of photosynthesis can be expressed
  as quantum yield (?).
• Quantum yield is number of photochemical product
  produced per photon absorbed.

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Noncyclic electron transport

• When electron transport is operating according to
  the figure above, electrons are continuously
  supplied from water and withdrawn as NADPH. This
  flow-through form of electron transport is known
  as noncyclic or linear electron transport.

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Cyclic electron transport

• Cyclic electron transport is referring to a
  condition when electrons from PSI is transported
  not to Fd-NADP-reductase but to a Fd-PQ
  reductase.

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Photophosphorylation

• The light-driven production of ATP by
  chloroplasts is known as photophosphorylation.

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How is ATP generated?
The light-driven accumulation of protons in the
lumen by oxidation of water and PQ-cytochrome
proton pump is the energy source of ATP
production.
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How cytb6f complex moves protons (H) across the
membrane

• The most widely accepted model for this question
  is known as the Q-cycle.

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Q-cycle (1)
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Q-cycle (2)
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ATP synthase complex
Thylakoid ATP synthase complex 400kDa, 9
subunit. CF1 (hydrophilic stromal part)
?3?3??? CF0 (transmembrane segment) I II III12IV
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Binding change mechanism of ATP synthesis by the
CF0-CF1 complex
O-site (open) available to bind ADP and
Pi L-site (loose) ADP and Pi are loosely
bound T-site (tight) nucleotide-binding
site Proton translocation ? conformation change ?
rotation of g ? interconversion of these sites
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Lateral heterogeneity

• Lateral heterogeneity is referring to the
  condition that two photosystems (PSI and PSII)
  are distributed unevenly.
• PSI is mainly located in the stromal membranes
  and PSII is in the granal membranes.
• ATP synthase is found mostly in stromal membrane.
• Cytochrome b6f complex is distributed evenly.

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Lateral heterogeneity
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Lateral heterogeneity

• Lateral heterogeneity is referring to the uneven
  distribution of PSI, PSII, and ATP synthase
  complexes on thylakoid membranes.

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Lateral heterogeneity
PSI/LHCI and ATP synthase
Cytb6f is uniformly distributed
PSII/LHCII
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Consequences of lateral heterogeneity

• The ratio between PSI and PSII is adjustable.
• Output of NADPH and ATP can be adjusted to meet
  cellular demands because non-cyclic and cyclic
  photophosphorylations can happen more or less
  simultaneously.

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Role of LHCII in photosynthesis

• LHCII contains more than half of the chlorophyll
  a and almost all of the chlorophyll b, however it
  is not directly involved in photochemical
  reduction.
• Functions of LHCII
• (1) increase the activity of PSII under
  conditions of low irradiance (shade plants)
• (2) regulate PSII activity when light condition
  fluctuates for a short period of time
  (phosphorylation/dephosphorylation)

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Shade plants

• Plants grown under shade have more thylakoids
  with large grana, therefore they have higher
  proportion of apressed thylakoids.
• Sun plants have less LHCII but with more
  cytochrome b6f complex, plastoquinone,
  plastocyanin, ferredoxin, and ATP synthease
  (CF0-CF1 complex).

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Phosphorylation/dephosphorylation of LHCII

• LHCII can be phosphorylated by a protein kinase.
  The phosphorylation causes LHCII becoming more
  negatively charged.
• Phosphorylated LHCII can be dephosphorylated by a
  protein phosphatase.

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Phosphorylation/dephosphorylation of LHCII

• Under high irradiance of light, PSII will be
  preferentially excited (state 2). The activation
  of PSII will result in accumulation of PQH2,
  which will activate (LHCII) protein kinase.

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Phosphorylation/dephosphorylation of LHCII

• The protein kinase is then phosphorylate LHCII.
• The phosphorylation makes LHCII becoming more
  negatively charged.

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Phosphorylation/dephosphorylation of LHCII

• LHCII moves to the stromal thylakoid because
  charge repulsion, making PSII antenna size
  smaller.
• Granal thylakoid also loosens due to lack of
  LHCII.

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Phosphorylation/dephosphorylation of LHCII

• Now PSI is preferentially excited (state 1).
• PQH2?, PQ?
• (LHCII) phosphatase is activated.

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Phosphorylation/dephosphorylation of LHCII

• Phosphatase dephosphyrylates LHCII and LHCII
  moves back to the granal side, which increase the
  antenna size of PSII.
• Granal membrane is stacked again.

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Figure 4.12
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LHCII and photoprotection

• PSII is the component of the thylakoid membrane
  that is most sensitive to excess light.
• Phosphorylation/dephosphorylation of LHCII will
  protect PSII from thermal damage due to excess
  light energy.
• Photodamage happens when excess light causes the
  oxidation of the D1 protein of PSII, which is
  slowly reversible to some extent.

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Carotenoid and photoprotection

• The principle carotene in most higher plants is
  b-carotene.
• Carotenoids serve two functions in
  photosynthesis light harvesting and
  photoprotection.

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Carotenoid and photoprotection

• Carotenoid-deficient mutant and
  norflurazon-treated plants (Norflurazon is an
  inhibitor of phytoene desaturase and subsequent
  blocking of carotenoid biosynthesis) are bleached
  in spite of their ability of chlorophyll
  biosynthesis is still functional.

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Carotenoid and photoprotection

• Carotenoids will trap and dissipate excess
  excitation energy before it reaches reaction
  center.
• If excess excitation energy (happens during
  periods of peak irradiance) reaches reaction
  center chlorophyll, the chance of 1O2 production
  (reactive oxygen species, ROS) will increase, and
  ROS will result in cell damage, even death.

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Xanthophylls
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Zeaxanthin and photoprotection

• Zeaxanthin can dissipate excess excitation energy
  as heat.

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Zeaxanthin is formed by xanthophyll cycle
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Xanthophyll cycle

• Under conditions of excess light, violaxanthin is
  enzymatically converted to zeaxanthin through
  de-epoxidation.
• De-epoxidation can also be induced by a low pH in
  the lumen, which also happens under high light
  conditions.
• Violaxanthin can also act as a light-harvesting
  carotenoid.

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Xanthophyll cycle

• Violaxanthin is a diepoxide. The de-epoxidation
  of it is progressing one by one, first producing
  antheraxanthin (monoepoxide), then zeaxanthin.
• Antheraxanthin and zeaxanthin will be converted
  back to violaxanthin in the dark by enzymatic
  actions.

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Xanthophyll cycle

• Both antheraxanthin and zeaxanthin can lose
  excess energy in the form of heat.
• However, neither of they can transfer their
  energy to chlorophyll because even when they are
  in excited states, their energy levels are still
  lower than antenna chlorophylls.

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Xanthophyll cycle

• Although they cannot pass their energy to antenna
  chlorophyll, antenna chlorophyll can transfer
  excess energy to them and dissipate it as heat.

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Xanthophyll cycle

• So xanthophyll cycle acts as a switch, generating
  antheraxanthin and zeaxanthin whenever
  dissipation of excess energy is required but
  removing the zeaxanthin under conditions of low
  irradiance.

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Potential value of xanthophyll cycle
Shade leaves Sun leaves
Xanthophyll content 13 32
Absorbed light used in photosynthesis 91 12
Light allocated to dissipation as heat 6 79
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Mehler reaction and Asada-Halliwell pathway

• Sometimes (about 510) O2 can react with
  electrons generated by PSI, producing superoxide
  radical (O2-). This is called Mehler reaction.
• Superoxide dismutase (SOD) will remove the O2-,
  producing H2O2 (peroxide). H2O2 is then reduced
  to water by ascorbate.

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Mehler reaction and Asada-Halliwell pathway

• Plant chloroplasts normally exhibit relatively
  high concentrations of ascorbate (0.51.0 mmol/mg
  of chlorophyll).
• This pathway is to prevent H2O2 react with O2-,
  producing OH(hydroxyl radical).

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Chlororespiratory pathway reducing O2 in the
dark
O2 H2O
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Chlororespiratory pathway

• Chlororespiartory pathway is probably have a role
  in photoprotection because it is not only
  operating in the dark.
• This pathway also operate in the light when
  organisms are exposed to excess irradiance.

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The D1 repair cycle
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The D1 repair cycle

• PSII reaction center exhibit an inherent lifetime
  because D1 polypeptide of PSII will be
  irreversibly damaged due to photo-oxidative
  damage after absorption of 105 to 107 photons.
• The life span for each D1 polypeptide of PSII
  reaction center is about 30 minutes.

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The D1 repair cycle
marked for degradation
psbA
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D1 polypeptide

• In addition to prone to photooxidation damage, D1
  polypeptide is also the binding site of many
  herbicides. Therefore it is also called herbicide
  binding protein).
• Herbicides belong to urea derivative and triazine
  groups inhibit photosynthesis by binding to QB
  site of D1 polypeptide, interrupting
  photochemical electron transport.

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Urea derivatives and triazines
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Triazines

• Some plants are resistant to triazines so it can
  be used as a selective herbicides.
• Corn roots contain an enzyme that degrade the
  herbicide. Cotton sequesters the herbicide in
  special glands.
• Some weeds also develop resistance toward this
  herbicides.

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Bipyridylium viologen dye herbicides

• This class of herbicides act by intercepting
  electrons on the reducing side of PSI, thus
  interrupting electron transport.
• After accepting electrons from PSI, they
  auto-oxidize and reduce oxygen to superoxide,
  which cause oxidative damage to plants.
• Herbicides in this class is also toxic to animal,
  therefore the usage is highly regulated.