VL 3

• balamcing electron ransfer PSI - PSII: State tramsition

• Nom Photochemical Quenching (NPQ)

• theory: always an even distribution of the excitation energy due to antenna movement

• plastiquinone redox state (PQ/PQH2) regulates the LHCII-kinase activity

• western blot analysis of phophorylated threonine

> STN7 is the LHC-Kinase

> changein the distribution of excitation energy

> change in fluorescence is not fully reversible

• not all changes depend on STN

• blue natve page of thylakoid membrane of Arabidopsis in state 2, solubilised with digitonin

  > seperation of PSI-LHCI-LHCII

> complex exist

• but how big?

• which subunits?

• PSII with and without LHCII

• free LHCII

• PSI-LHCI with and w/o LHCII

> prrofen: LHCII trimer is asociated with the PSI in State 2

> confirmed at higher solution

• PSII is surrounded by LHCII trimers (antenna complexes) mainly composed of Lhcb1, Lhcb2 and Lhcb3.

• Two LHCII trimer types are observed: trimer‑M (moderately bound, mobile; enriched in Lhcb1/3) and trimer‑S (strongly bound, immobile; enriched in Lhcb1/2).

• Under changing light, mobile LHCII (from trimer‑M) can dissociate from PSII and create a free pool.

• Some mobile LHCII can attach to PSI to form PSI–LHCI–LHCII supercomplexes, but not all LHCII that leaves PSII reaches PSI.

• Functional consequence: LHCII mobility enables dynamic redistribution of excitation energy (state transitions) and photoprotection.

• P-Lhcb2 bnds in the complex

• PSI & PSII have their own pool of LHCII

• PSII: seperation after phosphorylation (STN7)

  PSI: binding after phsphorylaton = state 2

• de-phosphorylation vice versa = state 1

• change in the distribution of excitation energy between the photosystems by adaptaion of the relative antenna size

• photoprotection is the state between starving and burning

• absorption of light cannot be regulated on the molecular level

• balace between effiient photosynthesis and avoidance of photo-oxidative detruction has to be kept

> excess excitation energy is dissipated as heat harmless

> ß-carotene or Non-Phoochemical Quenching (NPQ)

• all pathways are competing with each other

• the longer the excited single state (1Chl) is present, the higher the probability of triplet Chl (3Chl)

• 3Chl produces singlet oxygen and from there reactive ixygen specied (ROS) and has to be prevented

• direct removal of triplet Chl or singlet oxygen

• close to reaction center

• in PSII as last rescue

  > in CP43/CP47 (inner antennae), and additionally in the reaction center

• PAM (pulse amplitude modulated) - fuorescent measurement, so-calles quenching analysis

• the parts of the light energy that are coverted to heat or used for photochemistry can be calculated

  > quenching by pulsing

• NPQ: measure of heat dissipation

• under strong illumination photosynthesis becomes saturated and heat imission increases

• NPQ correlates with an increase in zeaxanthin concentration and a decrease in violaxanthin concentration = xantophyll cycle

• binding site in LHCII

• located between monomers of LHCII

• violaxanthin has to be de-epoxidised at both ends

• de-epoxidase: member of the lipocalin-family (barrel-like bnding domain for lipid f pigments)

• dimerisation at pH5 and association on the lumenal side of the thylakoid membrane

• present: LHCII, minor LHC and LHCI?

• theoretically: energy leel of Zea enables energy transfer from Chl a

• energy level of Zea in organic solvent enables energy transfer from Chl a

• gear shift (Chl+ + Zea > Chl + Zea* > Chl + Zea + heat)

  > Energy level in the proteins are not fitting

• still possible: other intermediates (different excited states than Zea*)

• yes, under NPQ conditions

• zeaxanthin is made in LHC and afzer excitation Chl a takes an electron temporarily from Zea

  > recombination (`giving back`) the electron under heat dissipation

• no

• present in CP24, CP26 and CP29 - NPQ reduction

> means that one of the central xantophylls has to be de-epoxidised

>although cation radicals might develop, nothing can be regulated

• de-epoxidanr: possible via lipocalin-like de-epoxidase

• energy level of zeaxanthin in vivo: gear shift not possible

• zeaxanthin radicals cations in minor LHC are possibly not involved

• lacking in Lhcb3, CP24 and CP29

• de-epoxidant: possible via lipocalin-like de-epoxidase

• energy level of zeaxanthin in vivo: gear shift not possible

• zeaxanthin racidal cations in minor LHC are possibly not involved

• aggregation of minor LHC/LHCII has to occur (CP24/CP29/LHCII)

• NPQ does not depend solely on Zea, LHC II conformational change as well as aggregation is needed, lutein quenche as well, meanhwhile new S* carotenoid state identified

• LHCII exists in two states: `quenched` or `fluorscing`

• neoxanthin can be twisted or straught

• lutein takes enegry from Chl a and transfers it to Chl a

• yes, because withour PsbS no NPQ!

• LHC:

• Chl a

  Chl b

• lutein

• neoxanthin

• violaxanthin

• monomer/trimer

• PsbS

• belongs to the LHC family

• 4 helices instead of 3

• no pigments

• dimer

• can be protonated on the lumenal surface

• changes its conformation in a pH-dependent manner

  > might be able to detect lumenal acidification

• Modell 1:

  • helps with stacking and the LHC association to PSII when un-protonated

  • when protonated: helps with unstacking and LHC dissociation from PSII

    > decrease in fluorescence (NPQ)

• Modell 2:

  • LHC forms a complex with PsbS (NPQ)

  • if Zea bound in addition = stronger NPQ

• correlates with increasing zeaxanthin content, zea+* can be proven in minor LHCs

• NPQ needs PsbS

• NPQ needs the formation of CP29-P24-LHCII supercomolexes

• also LHCII aggregates and is then quenched, S* state of carotenoids is formed

• psbS changes the association PSII-LHCII in a H-dependent manner

• psBs can be protonated and changes its conformation

• psbS increases NPQ by complex formation with LHCII

• CP29 zeaxanthin cation radical can be proven, but cannot be regulated

• in LHCII > Chl/Lut

• aggregates LHC are more strongly quenched > Car/Chl

• NPQ is very important and tus multilayered mechanism