Overview
(KEGG Pathway MAP00195)

All energy consumed by biological systems ultimately comes from light energy trapped in reduced carbon skeletons by the process of photosynthesis.

This simple net reaction is catalyzed by a very complex enzyme machinery in plants and photosynthetic bacteria. Photosynthesis traps light to convert photon energy into chemical energy in a process reminiscent of oxidative phosphorylation. This membrane bound process is referred to as light reaction and depends on an electron transfer chain powered by reducing equivalents extracted from water. The chemical energy produced - ATP and NADPH - is used for for the synthesis of carbohydrates by carbon fixation (Calvin cycle or dark reaction).

Chloroplasts

This chapter presents the plant photosynthetic apparatus. It is located in specialized organelles called chloroplasts with a complex membrane system that allows the separation of charges to convert electromagnetic energy (photons) into electrochemical energy (proton gradient) and eventually into chemical energy in the form of ATP (phosphoryltransfer potential) and NADPH (reducing power) and ultimately as carbohydrates, fats and amino acids..

Photosynthetic membranes and the light reaction

The light reaction is a membrane bound process. The photosynthetic membranes in plants contain two photosystems, photosystems I and II, or PS I and PS II. Photosystem I captures the energy of single photons by excitation of electrons in magnesium ions in green pigments called chlorophyll (chlorophyll a, C05306; see pathway map details below). The color of the chlorophylls indicates the absorption maxima of the system, i.e., the ability to capture light energy of photons at a specific wavelength. Photosystem I is coupled to an NADP⁺ - reducing enzyme to produce reducing power in the form of NADPH/H⁺. This reduction requires a steady supply of electrons and protons (reducing equivalents) These reducing equivalents are provided by a second photosystem which oxidizes water to molecular oxygen. Photosystem II uses the energy from a second photon to reduce plastoquinone QH₂. The reduced quinone are deoxidized by a cytochrome bf complex separating electrons from protons generating a proton gradient. The proton gradient is coupled to H⁺-ATPases similar to those in mitochondria synthesizing ATP from ADP + P_i.

Photosynthesis is not simply a reversal of oxidative degradation of carbohydrates and oxidative phosphorylation in mitochondria. The difference lies primarily in the source of high potential electrons. For oxidative phosphorylation the electrons come from hydrocarbon and carbohydrate structures, whereas in photosynthesis, the electrons have to be energized in the magnesium center of chlorophylls. The trick is to find a primary source of electrons to replenish the electrons extracted from chlorophylls by means of electromagnetic excitation. This primary source is H₂O. The energy potential of electrons in water is very low (water is not a reactive solvent) and it is a special metal ion complex (contains manganese) in PSII that has an even lower energy potential for electrons oxidizing water into the radical O^. and hydrid H^-, two very unstable, reactive intermediates. Photosystems II and I are coupled by the cytochrom bf complex and the electrons extracted from water by PS II (strong oxidant) and upon photon activation is transported by the electron transport chain via plastoquinone (weak reductant) to the cytochrome bf complex and from there to PS I via plastocyanin (weak oxidant) where it is activated by a photon into activated PS I  (strong reductant) which in turn is capable of reducing NADP⁺. The photosystems simply provide a means of increasing the high reduction potential necessary, using two photon activated steps, to push electrons from water to NADP⁺.

 

Photosystem II	Photosystem II is a multi subunit complex (10 subunits) with a molecular weight > 600kDa. It catalyzes the light driven electron transfer from H2O to plastoquinone (Q; oxidized form C02061). The net reaction for one photon (hn) is:                                 2Q + 2H2O Þ O2 + 2QH2 Plastoquinone exists in three different redox states; the fully oxidized Q (non charged), the half reduced/oxidized semiquinone form QH.-, and the fully reduced plastoquinol form QH2. The photosystem II protein complex contains three subcomplexes, the light harvesting complex II, the reaction center RC, and the oxygen evolving unit. The light harvesting complex II (LHC2) contains accessory pigments to capture photons. There are seven chlorophyll a, six chlorophyll b, and 2 carotinoid units in LHC2. The difference in the different pigments is their maximum of absorption, which broadens the spectrum of available sunlight to excite electrons. At the core of the complex lies the reaction center (RC) which is a dimer consisting of subunits D1 and D2, a special pair chlorophyll a (with an absorption maxima at 680nm; P680), and an electron transport chain across the thylakoid membrane consisting of pheophytin (with 2H+ instead of a Mg+ in the porphyrin ring) and two quinones, quinone A and quinone B (plastoquinone Q). Quinone A is tightly bound to the reaction center complex, whereas QB is able to dissociated in the fully reduced form QBH2 (Plastoquinol-1; reduced QB;  C02185). The electron, activated by a photon in the special pair at the lumenal side of the thylakoid membrane, is now located at the stromal side. The result is a charge separation across the thylakoid membrane which is now used by the membrane soluble plastoquinole which diffuses across the membrane back to the lumenal side where it is being oxidized by the cytochrome bf complex. It donates its electrons to the cytochroms and the protons to the thylakoid lumen aqueous compartment. Thus a proton gradient is established in this first half reaction between PS II and cytochrome bf complex. The oxygen evolving complex catalyzed the oxidation of water by a manganese complex as shown. Four electrons are sequentially extracted from 2 molecules of water producing molecular oxygen (O2) and 4 H+ that are used for the reduction of plastoquinone. The Mn-complex is in its S(0) state, the fully reduced state and can donate electrons to P680 in the reaction center. The S(4) state is the fully oxidized state capable of extracting electrons from water molecules. The extraction process needs two water molecules bound simultaneously to form one molecule of molecular oxygen to prevent the formation and release of free oxygen radicals that would damage the plant cell.
Cytochrome bf complex	The cytochrom bf complex essentially couples photosystem II and I and also provides the means of proton gradient formation by using cytochrom groups as redox centers in the electron transport chain thereby separating the electron/hydrogen equivalent into its electron and proton components. The electrons are transferred to photosystem I via plastocyanin and the protons are released into the thylakoid lumen of the chloroplast. The net reaction of the cytochrome bf complex is:                             QH2 + 2PC(Cu2+) Þ Q + 2PC(Cu+) + 2H+ The protein complex contains 4 subunits, a 33kDa cytochrom f, 23kDa cytochrome b563 (two hemes), a 20kDa Fe-S protein, and a 17kDa protein.
Photosystem I	Photosystem I is a membrane protein complex with 13 subunits and a molecular weight of > 800kDa. It catalyzes the net reaction:                             PC(Cu+) + ferredoxinox Þ PC(Cu2+) + ferredoxinred Again, the membrane protein complex promotes a charge separation transferring an electron from the lumenal to the stromal side of the thylakoid membrane. The energy for the transfer comes from light absorption by pigment P700, a special pair of chlorophyll a, and the subsequent electron transfer via quinones A0 and A1 (vitamin K1; reduced form C03313), a 4Fe-4S complex, and from there to ferredoxin. The enzyme ferredoxing-NADP+-reductase, a flavoprotein containing protein, reduces NADP+ to NADPH. Photosystem I can run in two modes. First, it promotes the reduction of NADP+ to provide reductive power for biosynthetic purposes. Second, photosystem I can switch to a cyclic operation where cytochrom bf complex serves as the electron acceptor instead of ferredoxin-NADP-reductase. The electron cycles from ferredoxin (Fd) to plastocyanin (PC) and back coupling to a proton transport for maximal ATP synthesis. The cyclic mode is stimulated when NADPH levels are high. P700 is the PS I special pair chlorophyll a which is photon activated (P700*) to drive the cycle.

Carbon fixation (dark reaction)  -  Calvin Cycle

The dark reaction of photosynthesis catalyzes the fixation of carbon dioxide into carbohydrates. This process is performed by the Calvin cycle which resembles the reversal of the pentose phosphate pathway. The fixation step is catalyzed by the enzyme RUBISCO (ribulose1,5biphosphate carboxylase/oxygenase):

                   Ribulose1,5biphosphate + CO₂ Þ 2 Glycerate-3-phosphate

The synthesis of D-Ribulose 1,5-bisphosphate (C01182) can be found in KEGG pathway maps MAP00630 (Glyoxylate and dicarboxylate metabolism) and MAP00710 (Carbon fixation).  The carboxylase/oxygenase enzymes catalyzing this carbon dioxide fixing reaction have E.C. numbers 2.7.1.19 (R) and  4.1.1.39 (R), respectively. Glycerate-3P is phosphorylated and reduced to glyceraldehyde-3-phosphate, a glycolysis/gluconeogenesis intermediate and, in a combination of transketolase and transaldolase reactions analogous to the pentose phosphate pathway, recycled to provide ribulose-5-phosphate. The latter can be further phosphorylated by ATP hydrolysis to form ribulose-1,5-biphosphate. The net energy balance of six rounds of the Calvin cycle to produce one mol of hexose is thus:

        6CO₂ + 18ATP +12NADPH + 12H₂O Þ C₆H₁₂0₆ + 18ADP + 18P_i + 12NADP⁺ + 6H⁺

The Calvin cycle's hexose product is fructose-6-phosphate which can be converted to glucose-6-phosphate by the gluconeogenetic pathway. Glucose-6-phosphate can be used for starch and cellulose synthesis (see section on glycogen metabolism).
  The Structure of Rubisco is known at high resolution and can be found at the protein data base (PDB entry for Rubisco L subunit (biological unit is an octamer) is 1A7J. Rubisco is a large enzyme complex on the stromal surface of the thylakoid membrane with 8 large subunits of 55kDa each (L subunits) and 8 small subunits of 13kDa each (S subunits). The L subunits contain both the catalytic and regulatory sites of the complex. The protein is activated by carbamate formation by RuBP carboxylase activase. A CO₂ unit is covalently linked to the epsilon amino group (e -NH₂) of a lysine residue (lysine-COO^-; carbamate) of an L subunit.

Control of Calvin cycle

Carbon dioxide fixation (dark reaction) occurs during the day time and is strictly dependent on the light reaction, i.e., the formation of reductive power as NADPH. It does not occur literally in the dark (at night). The metabolic activity at night instead is mainly shifted to carbohydrate oxidation, when plants consume oxygen rather than producing it. The control of the Calvin cycle therefore means the control of regulation between light and dark reaction. The enzymes of the Calvin cycle are sensitive to the proton concentration of the chloroplast stromal compartment. The optimum pH for Rubisco activity lies around pH 8. The proton pump driven during light absorption moves protons from the stroma to the lumenal side to the thylakoid membrane increasing the stromal pH. This activates Rubisco and thus the Calvin cycle. The proton level control is coupled to magnesium interaction (chelation) with carbamates. Magnesium stabilizes carbamate formation and thus activates Rubisco. The key enzymes fructose biphosphatase and sedoheptulose biphosphatase of the Calvin cycle are also under pH control. In addition, they are sensitive to the redox potential of the chloroplast stromal compartment. Biphosphatases are active in their reduced state only sensing the oxidation state of photosystem I. The mediator molecule is the small protein thioredoxin. Thioredoxin is coupled to ferredoxin by ferredoxin-thioredoxin reductase.   Thioredoxin also inhibits phosphofructokinase. In plant cells, light stimulates the Calvin cycle enzymes (PSI being the strong reductant) and inhibits glycolysis.

Photorespiration

Rubisco catalyzes a second reaction that also uses ribulose1,5biphosphate (RuBP) as substrate in the presence of molecular oxygen instead of CO₂. This process is known as photorespiration and yields phosphoglycolate (a C2 compound) and 3-phosphoglycerate (a Calvin cycle intermediate). Phosphoglycolate cannot be used by the Calvin cycle enzymes and two units are converted into 3-phosphoglycerate via a complex pathway via glyoxylate (in peroxyxysomes). Glyoxylate is transaminated to glycine. Two glycine molecules are combined to form serine (minus one carbon dioxide) which is oxidized to 3-phosophglycerate. Thus, photorespiration consumes molecular oxygen (in chloroplast) and generates carbon dioxide (in mitochondrion). Because this process is the result of the competition of molecular oxygen and carbon dioxide for Rubisco, photorespiration reduces the overall yield of photosynthesis (light plus dark reaction).

C4 plants

Not all plants show the same efficiency in photosynthetic carbon fixation. Certain plants produce high levels of CO₂ which helps reducing the energy wasting effect of photorespiration by competing with molecular oxygen for Rubisco. These so called C4 plants have a malate/pyruvate shuttle mechanism between mesophyl cells and bundle sheath cells to provide high carbon dioxide concentration in the latter. Mesophyl cells carboxylate phosphoenolpyruvate to oxaloacetate (C4), which is reduced to malate. Malate in turn is transported into the neighboring bundle sheath cells where it is decarboxylated to pyruvate. The released CO2 serves as substrate for Rubisco. Pyruvate is transported back into mesophyll cells and regenerated to phosphoenolpyruvate (see gluconeogenesis).

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