Ivar Virgin

Photoinhibition of Photosystem II. Inactivation, protein damage and turnover.

Even though light is the source of energy for photosynthesis, it can also be harmful to plants. Light-induced damage is targetted mainly to Photosystem II and leads to inactivation of electron transport and subsequent oxidative damage of the reaction centre, in particular to the D1 protein. Inactivation and protein damage can be induced by two different mechanisms, either from the acceptor side or from donor side of P680. The damaged D1 protein is triggered for degradation and digested by at least one serine-type proteinase that is tightly associated with the Photosystem II complex itself. The damaged Photosystem II complex dissociates from the light-harvesting antenna and migrates from appressed to non-appressed thylakoid regions where a new D1 protein is co-translationally inserted into the partially disassembled Photosystem II complex. D1 protein phosphorylation probably allows for coordinated biodegradation and biosynthesis of the D1 protein. After religation of cofactors and assembly of subunits, the repaired Photosystem II complex can again be found in the appressed membrane regions. Various protective mechanisms and an efficient repair cycle of Photosystem II allow plants to survive light stress.

Strong light photoinhibition of electrontransport in Photosystem II. Impairment of the function of the first quinone acceptor, QA

Abstract Electron paramagnetic resonance (EPR) spectroscopy has been applied in an investigation on the mechanism for photoinhibition of the electron transport in Photosystem II. The experiments were performed in vitro in thylakoid membranes and preparations of Photosystem-II-enriched membranes. Photoinhibition resulted in inhibition of the oxygen evolution and EPR measurements of the S 2 state multiline EPR signal show that its induction by illumination at 198 K was decreased with the same kinetics as the oxygen evolution. Further EPR measurements show that the reduction of Q A was inhibited with the same kinetics as the oxygen evolution. The amount of photoreducible pheophytin was estimated from photoaccumulation experiments under reducing conditions and the results show that the primary charge separation reaction was inhibited much slower than the oxygen evolution or the reduction of Q A . These results indicate that photoinhibition inhibits the electron transfer between pheophytin and Q A probably by impairment of the function of Q A . In the inhibited centers the primary charge separation reaction is still operational. It is suggested that the event leading to photoinhibition of the electron transport is the double reduction of Q A which then leaves its site. Photoinhibition also results in rapid oxidation of cytochrome b -559 and a change of cytochrome b -559 from its high potential form to its low potential form. The reaction is quantitative and proceeds with the same kinetics as the inhibition of oxygen evolution. The potential shift of cytochrome b -559 suggests that photoinhibition induces early conformational changes in Photosystem II.

Photosystem II disorganization and manganese release after photoinhibition of isolated spinach thylakoid membranes

The activity, the protein content and the manganese properties of photosystem II have been compared after photoinhibition of isolated thylakoid membranes. The results show a concomitant disappearance of the oxygen evolving activity and the ability to form the S2‐state multiline EPR signal. The D1‐protein is degraded in a subsequent event which closely correlates to release of manganese from the thylakoid membranes.

Changes in the organization of Photosystem II following light-induced D1-protein degradation

Abstract The composition and organization of photosystem II was studied in thylakoid membranes and subfractions which had been subjected to photoinhibitory light conditions. The results show that D1-protein degradation can occur in vitro leading to a 50–60% loss of the protein. Apart from the D2-protein, which shows a limited decline, there was no loss of any other photosystem II proteins. The D1-protein degradation induced several changes in the organization of the photosystem II complex. Using inside-out thylakoid vesicles we demonstrate that concomitant with the D1-protein degradation there is a release of the extrinsic 33, 23 and 16 kDa proteins from the inner thylakoid surface into the lumenal space. In addition, there is a release of four manganese ions per D1-protein degraded. The correlation between the D1-protein degradation and the release of manganese is also seen in inside-out thylakoid vesicles that have been CaCl2-washed to remove the three extrinsic proteins prior to photoinhibitory illumination. Subfractionation of thylakoids subsequent to photoinhibitory treatment suggests a migration of certain photosystem II subunits from the appressed to the non-appressed thylakoid regions following D1-protein degradation. The photosystem II subunits showed an individual migration behaviour, suggesting a disassembly of the photosystem II core. Our data suggest that repair of photodamaged photosystem II involves, apart from reinsertion of new D1-proteins, reassembly of the photosystem II complex including lateral movement of proteins between the two thylakoid regions and religation of the manganese cluster.

Photodamage to photosystem II : Primary and secondary events

Abstract High light stress results in a reduction in the photosynthetic capacity of plants. This photoinhibition is targeted to photosystem II and seems to be an inevitable consequence of the complicated redox chemistry involved in the light-driven water-plastoquinone oxidoreduction reaction. Photoinactivation leads to irreversible damage of the reaction centre of photosystem II, in particular the D1 protein. In this paper, we present evidence which indicates that the inhibition of electron transport can be initiated at both the acceptor and donor sides of photosystem II. In the former case, there is a sequential array of events leading to overreduction of the primary quinone acceptor Q A . This, in turn, leads to the formation of a P 680 chlorophyll triplet. The donor-side inhibition is thought to involve the accumulation of the highly oxidizing species P + 680 and Tyr + z . These species will lead to oxidative damage of the D1 protein, triggering it for degradation. During acceptor-side inhibition, the protein is damaged by singlet oxygen formed via a reaction between P 680 triplet and oxygen. Damaged D1 protein is degraded by a serine protease located in the photosystem II complex. Preliminary inhibition and binding studies using the classical serine protease inhibitor diisopropylfluorophosphate suggest that the active serine is located on a 43 kDa polypeptide. Several D1 protein digestion fragments have been identified and a cleavage site between the stromally exposed loop connecting transmembrane helices IV and V is suggested.

Light‐induced D1‐protein degradation in isolated photosystem II core complexes

Photoinhibitory illumination of isolated oxygen evolving photosystem II core complexes results in a substantial degradation of the Dl‐protein which is accompanied by the appearance of high amounts of at least 4 different degradation products. It is suggested that the degradation is due to a protease that is an integral part of the photosystem II complex.

Light-induced D1 protein degradation is catalyzed by a serine-type protease.

Light‐induced degradation of the D1 protein in isolated spinach photosystem II core preparations was studied after addition of various protease inhibitors. The degradation was selectively inhibited by several serine protease inhibitors in particular diisopropylfluorophosphate. The results demonstrate that the D1 protein is degraded by a serine‐type of proteolytic activity that is an integral part of photosystem II.

Molecular mechanisms behind light-induced inhibition of photosystem II electron transport and degradation of reaction centre polypeptides

Excessively high light intensities are damaging to the photosynthetic apparatus. Photosynthetic organisms are therefore faced with the problem of maintaining sufficient excitation power under limiting light conditions whilst avoiding photodamage under high light conditions. The primary target for this photodamage is Photosystem II (PSII), which undergoes inhibition of its electron transport followed by irreversible damage to and degradation of the D~ and D 2 reaction centre polypeptides. This degradation is the initial event in a repair cycle of photodamaged complexes and is followed by protein synthesis and reassembly of functional PSII. This report will deal with recent information relevant to the molecular mechanisms of PSII electron transport impairment and of protein degradation, particularly the D I protein, and the chemical link between these two events. Data will be presented in support of the following sequence of reactions (Fig. 1), which take place at PSII under high-light stress: (i) overreduction of the acceptor side, leading to the formation of stably reduced QA species; (ii) these events facilitate the formation of chlorophyll triplets which react with molecular oxygen to form singlet oxygen; (iii) this highly reactive and damaging species will oxidize pigments and/or amino-acid residues leading to irreversible damage to the reaction centre, mainly to the D l protein; (iv) the oxidative damage induces a conformational change in the D~ protein which triggers it for

Structure of the Genes for LHC-II in Scots Pine (Pinus sylvestris L.)

Pines are in contrast to angiosperms capable of forming chlorophyll and probably all photosynthesis-associated proteins in the dark. However, the thylakoids of dark-grown (“etiolated”) pine seedlings differ from light-treated material in at least two ways: We and others has shown that the chlorophyll a/b ratio is higher, which indicates that the antenna size is smaller. PSII activity is also affected, there is a limitation on the water-splitting side of PSII in the dark-grown seedlings. In order to study the structure and regulation of pine photosynthetic genes, we wanted to isolate cDNA clones encoding LHC-II polypeptides from Scots pine.

Consequences of Light Induced D1-Protein Degradation on Thylakoid Membrane Organization

Photosystem II (PS II) of higher plant possess unique properties with respect to function, organization and protein turnover. It is a multisubunit protein complex which is composed of at least 20 different polypeptides (1). The two reaction center polypeptides, designated D1 and D2, appear to carry all the redox components necessary for the primary photochemistry of PS II (2) and possibly also the Mn (3). The great majority of the PS II units is located in the appressed thylakoid regions in association with its chlorophyll a/b antenna (4). PS II has a central catalytic role, but it also plays a central role in the long and short term acclimation of the photosynthetic apparatus. It is also the target for the photoinhibition process which leads to impaired electron transport capacity and the subsequent breakdown of the two reaction center subunits, in particular the D1-protein.