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PROBLEM OF ORIGIN OF LIFE International
Conference in Honor of 120th Birth Anniversary of
acad. A.I. Oparin Karapetyan N.V.
A.N. Bach Institute of Biochemistry RAS, Moscow
How cyanobactria managed to survive under
intense solar radiation billions years ago
Photoprotection mechanisms
September 26, 2014
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Acad. A.I. Oparin was elected as the first
President of ISSOL (photo was taken in Pont-
á-Mousson, France 1970)
.
You are our Pope, we are your monks!
?? ??? ???????, ?? ???? ?????!
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Acad. A.I. Oparin was the Director of A.N. Bach
Institute of Biochemistry for 1946-1980 Many
laboratories of our Institute have been involved
in study of Origin and Evolution of Life. My
contribution Photoprotecton mechanisms
against photodestruction by excess absorbed
energy in cyanobacteria. We have found two
mechanisms of photoprotection in
cyanobacteria 1.Carotenoid-less
non-photochemical quenching by Photosystem I
2.Carotenoid-induced non-photochemical quenching
of Phycobilisomes
.
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Cyanobacteria, the first photosynthetic
organisms, have originated about 2.5-3 Gyrs ago
in conditions of intense UV and VIS light at the
absence of ozone layer. Irradiance conditions on
the Earth surface NOW on the width of equator
UV-C (190-280 nm) - does not penetrate the
ozone layer UV-B (280-320 nm) - 7-8 W m-2 UV-A
(320-400 nm) - 45-50 W m-2 (generates singlet
oxygen) VIS light (400-700 nm) - 1100 W m-2 To
be protected against intense solar light and UV,
cyanobacteria were habituated in deep ocean
waters or in hydrothermal sources.

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Oxygenic photosynthesis
Photosynthesis is optimal under the balance of
the activity and stability of the photosynthetic
apparatus. Over-excitation of antenna Chls
generates reactive oxygen species that destroy
the photosynthetic apparatus. Dissipation (or
quenching) of excess absorbed energy protects
against photodestruction.
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• Carotenoid-less non-photochemical quenching by
  Photosystem I
• PSI complex exists in cyanobacteria as a trimer,
  in plants as a monomer.

3.4Å structure of PSI monomer of P.
sativum Amunts et al., Nature (2007)
2.5Å structure of PSI trimer of Th.
elongatus Jordan et al., Nature (2001)
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• Organization of Chlorophyll (Chl) antenna in
  cyanobacteria
• Chls in cyanobacteria are located only in core
  antenna of PSI and PSII since cyanobacteria are
  deficient in Chl-containing Lhca.
• Cyanobacteria are highly enriched with PSI
  PSI/PSII ratio is 3-5.
• Thus main part of Chls (90) in cyanobacteria is
  located in PSI.
• About 90 of antenna Chls in PSI of cyanobacteria
  belong to bulk while 10 of antenna Chls belong
  to long-wavelength Chls (LWC).
• The origin of LWC and the role in PSI was not
  clear.
• We have studied the role of the red-most LWC in
  energy balance and in energy dissipation in the
  cyanobacterium Arthrospira platensis
• Some information about LWC of PSI in
  cyanobacteria.

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LWC in PSI core antenna of cyanobacteria and
plants (GobetsKarapetyan et al., Biophys. J.
2001)
6 K
Gaussian deconvolution of 5 K absorption spectrum
of PSI trimers of A. platensis LWC740 (F760)
3 LWC708 (F730) 7 (Schlodder,.Karapetyan et
al., BBA 2005)
730 nm
290 K
trimer
740
708
monomer
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Spectral characteristics of LWC in PSI trimers
and monomers of A. platensis and Th. elongatus
amount of Chl molecules - in parenthesis
(Karapetyan et al., FEBS Lett. 1999)
Cyanobacteria Absorbance bands Fluorescence ?max with P700 red. Fluorescence ?max with P700 ox.
A. platensis trimers 708 (7) 740 (3) 727 760 726
A. platensis monomers 708 (7) 727 726
Th. elongatus trimers 708 (4) 719 (4) 730 741 732
Th. elongatus monomers 708 (4) 719 (2) 730 728
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Fluorescence DAS (decay associated
spectra) LWC delay the energy equilibration in
core antenna and trapping by P700 it is
dependent on spectral properties of LWC 35 ps in
PSI trimers of Th. elongatus - C 37 ps in PSI
monomers of A. platensis - D 50 ps in PSI trimers
of A. platensis - E. (Gobets,.. Karapetyan et
al., Biophys. J. 2001)
trimer
monomer
trimer
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P700 efficiently quenches F760 of PSI trimers of
A. platensis and F735 of PSI trimers of Th.
elongatus (Schlodder Karapetyan, BBA 2011)
PSI trimers
PSI monomers
P700AoA1-FxFA-FB-
A. platensis
760
P700AoA1FXFAFB
Th. elongatus

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• Energy transfer in PSI antenna depends on redox
  state of the cofactors of the
• PSI Rection Center (RC)
• open RC charge separation
• Chl ? P700A0A1FX ? P700Ao- A1FX
• closed RC dissipation of absorbed energy
• Chl ? P700A0A1FX or Chl ? 3P700A0A1-FX
• P700 is involved in charge separation
• P700 or 3P700 are involved in energy
  dissipation

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Origin of LWC interaction of Chl molecules on
the surface of various PSI monomers is forming
the red-most LWC (F760) in PSI trimers of A.
platensis (Karapetyan et al., Photosynth. Res.
1999)
Time-course of F760 quenching and P700 formation
in PSI trimers of A. platensis at 77K
Non-linear dependence of F760 on P700 amount in
PSI trimers of A. platensis indicates on energy
exchange between PSI monomers within trimer
PSI trimer of Th. elongatus (Jordan et al., 2001)
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Localization of LWC in PSI antenna of Th.
elongatus trimer 719 (F741) - 4 Chls 708
(F732) - 4 Chls monomer 719 (F730) - 2
Chls 708 (F728) - 4 Chls
Chl719 (F741) might be B7/A32/A31 Chl719 is
not B31/B32/B33 3 Chls, big distance to P700
(50Å) Candidates for Chl708 (F732) are B38/B37,
?38/A39, B18/B19 or A16/A17/A25 (strong coupling
between Chls, dig distance to P700). Chl715
(F734) B24/B25 or A26/A27
F741
F734
F734
F732
SchlodderKarapetyan, BBA (2012)
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Localization of LWC in PSI complexes of A.
platensis PSI trimer 740 (F760) - 3 Chl
708 (F727) - 7 Chl PSI monomer 708 (F726) 7
Chl (three different aggregates).
Chl740 (F760 ) might be A31/A32/B7 on lumenal
side close to trimerization point, time of
energy transfer to P700 is 110 ps, dipol is
oriented parallel to membrane
Chl708(F727)B38/B37, A38/A39 B18/B19 or
A25/A16/A17 Distance between Chl740 and
Chl708 Chl740
Chl708 A32/A31/B7 to B38/B37 22Å A32/A31/B7
to A25/A16/A17 48Å A32/A31/B7 to ?38/A39
57Å A32/A31/B7 to B18/B19 52Å
F760
F727
or F727
SchlodderKarapetyan, BBA (2012)
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Different orientation of Chls in various LWC730
of PSI antenna in A. platensis SMS
data Fluorescence spectra of a single PSI trimer
of as a function of the orientation of polarizer
in front of the spectrograph Chls in F730
polarized differently since 2-3 different
emitters form this LWC. Chls in F760 are
polarized equally. (Brecht,.Karapetyan BBA
2012)
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Scheme of energy migration in antenna of PSI
trimers of A. platensis No interaction of some
LWC708 and LWC740 at cryogenic temperatures -
big distance between F760 (?31-A32-B7) and LWC726
(different complexes) - different orientation of
the transient dipole moments in LWC708
(Karapetyan et al., Biochemistry-Moscow 2014)
Bulk Chl
P700
LWC708 F726
LWC708 F726
LWC740 F760
?31-A32-B7
P700
heat
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1. Conclusions PSI-induced energy dissipation in
cyanobacteria 1. LWC delay the energy
equilibration and trapping in PSI core antenna.
LWC function as terminal acceptors of excitation
like P700 and transfer uphill energy to P700.
2. P700 quenches the LWC fluorescence of PSI
trimers and monomers of A. platensis and Th.
elongatus but with different efficiency. 3.
LWC740 (F760) in PSI of A. platensis may
correspond to peripherally localized A31/A32/B7
trimeric aggregate. Localization of LWC719 in PSI
of Th. elongatus may differ since aggregate
contains 4 Chls.
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2. Caroteboid-induced NPQ of Phycobilisomes (PBS)
fluorescence in cyanobacteria PBS are the main
light-harvesting complex in cyanobacteria
Structure of Phycobilisomes, interaction with
Photosystems
PBS
PSII
PSI
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In 2004 we have found that illumination by
blue-green light of Synechocystis cells quenches
the fluoresence of PBS at 660 nm quenching is
reversible in dark (Rakhimberdieva et al., FEBS
Lett. 2004).
APC
dark (non-quenched)
after BL (quenched)
Action spectrum of quenching
Quenching decreases PBS fluorescence at 660 nm
(exc. 580 nm)
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Photoprotective dissipation of energy in
cyanobacteria. 1. PBS is the quenching target,
carotenoid is photosensitizer (Rakhimberdieva et
al., 2004) 2. Quenching - only at physiological
temperatures (Rakhimberdieva et al., 2004,
2007) 3. Quenching is ?pH independent
(Rakhimberdieva et al., 2006 Wilson et al.,
2006) 4.OCP-red (OCP) may be fluorescence
quencher (Wilson et al., 2006, 2008). Main
strategy to reveal the mechanism of quenching -
comparison of the activity of PSI and PSII in
Synechoystis mutant cells in non-quenched and
quenched states. PSI activity was measured for
PSII-less mutant, PSII activity - for PSI-less
mutant.
Orange Carotenoid-binding protein (OCP)
non-quenched
OCP (35 kDa) from A. maxima - two-domain
homodimer containing 3-hydroxiechinenone(Kerfeld
et al., 2003)
quenched
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down regulation of photosynthesis
Quantum efficiency of PBS absorption in
Synechocystis cells in quenched state drops by
about 40 (P700 photooxidation and PSII
fluorescence induction). OCP-triggered energy
dissipation in PBS of Synechocystis diverts
excitation away from both RC (Rakhimberdieva et
al., BBA 2010).
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BL-induced quenching takes place even at the
absence of PSI and PSII (Rakhimberdieva et
al., FEBS Lett. 2011)
Fluorescence quenching spectra at 77 K and RT
(top) and the second derivative of quenching
spectrum at RT (down).
77K fluorescence spectra (exc. 570 nm) of WT and
PSI/PSII-less mutant
77K
288 ?
77 ?
288 ?
680
660
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Light saturation curves of quenching centre
formation
BL
Kuzminov.. Karapetyan BBA 2012
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• 2. Conclusions on OCP-induced NPQ
• 1. Carotenoid is photosensitizer of PBS
  quenching, APC is a target of OCP-induced
  fluorescence quenching in Synechocystis cells.
• 2. OCP-induced quenching of APC fluorescence in
  Synechocystis cells diverts excitation energy
  from PBS to PSI and PSII reaction centres
  decreasing the energy flow from PBS.
• 3. Excitation of carotenoid in Synechocystis
  induces the multistep OCP transformation as
  sensitizer and as quencher.

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Thanks to colleagues Rakhimberdieva M.G.
A.N. Bach Institute of Biochemistry RAS,
Moscow Shubin V.V. Bolychevtseva Y.V Terekhova
I.V. Elanskaya I.V. Biology Faculty,
Genetics Dep., MSU Kuzminov F.I.
Physics Faculty, Dep. of Non-linear Fluorimetry,
MSU Schlodder E. Max-Volmer
Laboratorium, Technical University Berlin,
Germany Rögner M. Plant Biochemistry
Dep., Ruhr-University-Bochum, Germany Vermaas
W.F.J. School of Life Sciences, Arizona
State University, Tempe, USA