Energy transfer and charge separation in the PS 2 core complex

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Energy transfer and charge separation in the PS
2 core complex
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Experimental setup
800 nm lightTisapphire oscillator
amplifier Hurricane (Spectra Physics)
Visible lightNon-collinear Optical
Parametric Amplifier (second harmonic generator)
1 KHz 800 nm 0.8 mJ 80-90 fs
350 mJ
Delay 30 mm 100 fs
400-800 nm 5mJ, 10-30 fs
1150-2600 nm
IR1TOPAS (OPA)
MIDIR lightDifference frequency generator
450 mJ
2.4-11mm 3 - 1.5 mJ D ?200 cm-1
PROBE
PUMP
MIR window 200 cm-1detect between 1000 and 3500
cm-1 Sample is in moving CaF2 cell, Lissajous
scanner, Noise 10-5 OD in 1 minute
Spectrograph
SAMPLE
MCT
PC
preamplifier
IntegrateHold 16-bit ADC
pumped
unpumped
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CP43
CP47
RC
Figure 1. Pump-probe measurements in the mid-IR
upon excitation at 681 nm, Pexc 250 nJ. A)
Time traces detected at 1657, 1686 and 1711 cm-1.
The scale is linear up to3 ps and logarithmic
thereafter. The solid line through the data
points is a fit with time constants of 3 ps, 27
ps, 200 ps and 2.5 ns, the very fast component
that follows the IRF is also included. B)
Evolution Associated Difference Spectra
resulting from sequential analysis of the data
using a model with increasing lifetimes.
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• RC thermodynamic model plus 2 antennastates
• Use CP43 and CP47 spectra as input
• Include annihilation

Simultaneous fit of IR data and Streak-emission
data (Andrizhiyevskayaet al) of closed PS2 cores
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What are the energy transfer timescales in cores
of PSII?
keff 30-40 ps
CP47 17 Chls
1 ps
gt20A kgt2-3ps
RC
CP43 14 Chls
1 ps
gt20A kgt2-3ps
keff 30-40 ps
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The Major Organization Principles

• High pigmentprotein ratio and as a consequence
  ultrafast (lt 1 ps) energy transfer times.

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The Major Organization Principles

• High pigmentprotein ratio and as a consequence
  ultrafast (lt 1 ps) energy transfer times.
• Multiple Pathways for Energy Transfer and Many
  Entries Into the Reaction Center

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Multiple Pathways for Energy Transfer and Many
Entries into the Reaction Center
Chls at 3-4 nm transfer excitations in 10 ps!!!
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The Major Organization Principles

• High pigmentprotein ratio and as a consequence
  ultrafast (lt 1 ps) energy transfer times.
• Multiple Pathways for Energy Transfer and Many
  Entries Into the Reaction Center.
• Long Distance Energy Transfer vs. Short Distance
  Electron Transfer

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The RC-LH gap Förster vs. Marcus
Chlorophylls at a distance of 3-4 nm transfer
excitations in 10-30 ps.
And electrons in 1 second!!!!!!
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The Major Organization Principles

• High pigmentprotein ratio and as a consequence
  ultrafast (lt 1 ps) energy transfer times.
• Multiple Pathways for Energy Transfer and Many
  Entries into the Reaction Center.
• Long Distance Energy Transfer vs. Short Distance
  Electron Transfer.
• Long lifetimes in the absence of Reaction
  Centers

LH1/2 1 ns CP47 4 ns LHCII 2 ns,
etc Compare Chl a aggregates lt 20 ps!!!!!
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The Major Organization Principles

• High pigmentprotein ratio and as a consequence
  ultrafast (lt 1 ps) energy transfer times.
• Multiple pathways for energy transfer
• Many Entries into the Reaction Center
• Long lifetimes in the absence of Reaction
  Centers
• Multiple pigments-gtSpectrally broad
• Photoprotection In LHCII all chlorophyll
  triplets are transferred to carotenoid
  triplets!!!!
• Idem in LH1/2 and PS1

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The LHCII monomer
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What has been optimized?

• High pigmentprotein ratio!!!
• Long lifetimes in the absence of Reaction
  Centers
• Not the trapping time as such (lt 200 ps).
• Not the energy transfer time100 fs-10 ps!!!
• Rings, allowing sufficiently fast energy
  transfer into the RC, and sufficiently slow
  electron transfer into the antenna.
• Funnels????
• Multiple pigments-gtSpectrally broad ??????? EET
  efficiency??? Carotenoids30-90!!!
• Photoprotection chlorophyll triplets are
  transferred to carotenoid triplets.
• Regulation mechanisms

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• Excitation Energy Transfer
• and
• the Molecular Mechanism
• of
• Non-Photochemical Quenching

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The Efficiency of Photosynthesis

• The net reaction of photosynthesis is
• 2H2O CO2 gtCH2O O2H2O
• DG0485kJ/Mol
• For this reaction to turnover once requires 2x4
  light-events or 1800 kJ/Mol (assuming 500 nm
  photons)
• The maximum efficiency is about 10-15
• Biomass storage by plants is about 1

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Plant photosynthesis is not efficient
Gust, Kramer, Moore, Moore Vermaas, MRS
Bulletin, 2008, in press
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Use of the energy of light by plants

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Non-Photochemical Quenching in PS2 Transition
to a Dissipative State
Weak light
PS2RC
LHC
Fluorescence
Charge Separations
Strong light
PS2RC
LHC
Fluorescence
Charge Separations
3Chl, 1O2
Dissipation
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NON PHOTOCHEMICAL QUENCHING
Excess light illumination
The excess energy is thermally dissipated through
the quenching of the excited state of chlorophyll
PSII
Crucial steps
Activation of the xanthophyll cycle violaxanthin
gt zexanthin
Protonation of the PsbS protein
Direct quenching by zeaxanthin
Aggregation model for NPQ
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PS2 regulates the flow of energy towards the
reaction center
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LHCII AGGREGATES
AGGREGATION
-

-

CHL excited state lifetime
THE DEGREE OF AGGREGATION CAN BE CONTROLLED
THE QUENCHING CAN BE INDUCED IN THE ABSENCE OF
XANTHOPHYLL CYCLE CAROTENOIDS
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Ruban et al Figure 1
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THE EXPERIMENT
LHCII in three aggregation states, i.e. in three
quenching states Sample 1 least quenched

Sample 2 mildly quenched

Sample 3 strongly quenched
Probe (450-730 nm) at time delays (0-1 ns)
Excitation 675 nm (Chl Qy state) 40kHz
10nj/pulse
DATA ANALYSIS
Global analysis sequential model of increasing
lifetimes (EADS)
k1
k2
k3
A
B
C
A
In order to extract the spectrum of pure
molecular states a target kinetic model was
also applied to obtain species associated
difference spectra (SADS)
OBJECTIVES
Are carotenoids involved in the quenching process
?
If so, does the quenching proceed via energy or
electron transfer?
van Stokkum, I. H. M., Larsen, D. S. van
Grondelle, R. (2004) Biochimica Et Biophysica
Acta 1657, 82-104.
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TRANSIENT ABSORPTION SPECTROSCOPY
OD(?) -log I(?)/I0(?)
Excited state absorption
?OD(?,t) OD(?,t)pump on- OD(?,t) pump off
Ground state bleaching
Stimulated emission
Detector diode array
40 kHz, 7.5 µJ, 800 nm, 50 fs
475- 700 nm
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TiSapphire laser system
Optical Parametric amplifier
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Probe light 320-1100 nm
sample
NLM
L
L
Delay line (1ns)
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EADS NORMALIZED IN THE CHL Qy
REGION
Car bleach region
Car exc. state abs region
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Ruban et al Figure 3
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Ruban et al. Figure 2
Unquenched
Quenched
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Ruban et al Figure 3