Lesson 4: Photosynthesis - Processes

Transcription

1 Published on Agron 501 ( Home > Lesson 4 Lesson 4: Photosynthesis - Processes Developed by: By R. Shibles, D. Muenchrath and A. D. Knapp About 95% of plant dry matter is organic material produced by photosynthesis, a process unique to green plants and a few other organisms algae for example. Photosynthesis is responsible, directly or indirectly, forall the food and feed humans and other animals need to live. In fact, when you consider the source of fossil fuels, photosynthesis accounts for much of the energy we use today. Through photosynthesis (literally "building by use of light") plants construct themselves from carbon dioxide and water using solar energy to drive the process. The net reaction, where (CH 2 O) represents carbohydrate (a hydrated carbon), is written: Equation 4.1 Crop physiologists consider photosynthesis as three integrated processes: 1. The light reactions solar energy is captured and converted to chemical energy, 2. A diffusion process CO 2 moves from the atmosphere through the leaf into the chloroplast, and 3. A two step assimilation process CO 2 in the chloroplast is fixed and the resulting compound is reduced to form carbohydrate using the chemical energy from captured sunlight. Because photosynthesis is fundamental to yield, crop management techniques are designed to minimize any stress that would limit photosynthesis. In contrast, some herbicide families function by interfering with photosynthetic processes. Objectives 1. Describe the mechanism of solar energy capture and conversion to chemical energy in chloroplasts. 2. Explain the mechanisms of carbon diffusion to the site of carboxylation. 3. Understand carbon assimilation and carbohydrate production by chloroplasts. 4. Describe photorespiration and its impact on C3 and C4 plants. 5. Distinguish mechanistically between the three major types of CO 2 assimilation: C3, C4, and CAM. 6. Identify how and why the three photosynthetic types differ in terms of productivity and ecological adaption. Before you begin, listen to the instructor's comments on the key concepts in this Module.

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3 The Light Reactions The Nature of Solar Energy Solar energy is transmitted in waves as bundles of energy called photons, or quanta. Light capable of energizing photosynthesis is called photosynthetically active radiation (PAR) and encompasses wavelengths in the visible spectrum between 380 and 720 nm (Fig. 4.1). Because it occurs in the visible portion of the solar spectrum, PAR often is referred to casually as light. The spectrum for the perception of light by the human eye, however, differs from that of PAR for photosynthetic activity. Fig. 4.1 PAR and visible light portions of the electromagnetic spectrum. The quantity of PAR available for photosynthesis at a leaf surface is the Photosynthetic Photon Flux Density (PPFD). Some refer to this value as the "light intensity." Table 4.1 Comparative Solar Radiation Terminologies Radiation Name Wavelengths encompassed, λ Quantity Name Units of Quantity Maximum flux* 2200 light nm illuminance lux 14,000 µmol PAR nm PPFD photons m -2 s -1 radiant energy 0.1nm - 1.0m irradiance W m * at solar noon on a clear, summer day

4 Chloroplast Organization and Structure Photosynthesis occurs in a cellular organelle, the chloroplast (Fig. 4.2). Inside the chloroplast is a liquid matrix, the stroma. Suspended within the stroma is a system of flattened, sac-like membranous structures, the thylakoids. An individual thylakoid consists of two membranes enclosing an internal space, the lumen. Thylakoids occur in stacked groups, called grana, or as single stromal thylakoids that connect grana. The capture of PAR and its conversion to chemical energy, i.e. transduction, occurs within the thylakoids. Fig. 4.2 Photomicrograph of a sunflower chloroplast. Courtesy of Dr. Harry T. Horner, Department of Botany, Iowa State University, Ames, Iowa, U.S.A. The interactive activity Study Question 4.1 can only be completed online.

5 PAR Capture and Transduction to Chemical Energy Light energy is captured by various chlorophyll pigments and through the actions of photosystem I and photosystem II electrons and protons are stripped from the hydrogen atoms of water and separated. The electrons and protons so formed are separated by the thylakoid membrane to generate an electrochemical gradient that allows for the formation of two compounds. NADPH provides reducing equivalents for the reduction of CO 2 to CH 2 O. And ATP, a well known currency of energy in biological systems, is generated by proton mediated ATPases. Since the formation of ATP, in this case, is basically driven by light energy its formation is called photophosphorylation. Both the NADPH and ATP accumulate in the stroma of the chloroplast. The photosynthetic process of photophosphorylation occurs in two energy capture and conversion systems: Photosystem I (PSI) stationed in stromal thylakoids, and Photosystem II (PSII) located in granal thylakoids.(fig. 4.3) The spatial arrangement of PSI and PSII may be important for optimum distribution of photons between the two photosystems. Efficiency of energy conversion is greatest when the light absorbed by PSII and PSI is balanced so that both photosystems can function. Fig. 4.3 Schematic of a chloroplast with its thylakoids: granal (stacked) and stromal (unstacked area). (Adapted from Buchanan et al., 2000) Fig. 4.4 Close up diagram showing detail of chloroplast thylakoid systems. (Adapted from Buchanan et al., 2000)

6 The photosystems are linked by a series of electron carriers that form an 'electron transport chain'. These carrier molecules in the thylakoid membrane accept and release electrons based on their relative redox potentials. The arrangement of the photosystems and electron transport chain are conceptualized as the "Z-scheme" in the figure below. Z-scheme "Z-scheme" electron transport chain of PSII and PSI. Scale on left shows energy potential corresponding to the midpoint of carrier redox potential. Keep the organization of this Z-scheme in mind as we examine the sequence of reactions involving light harvest and energy conversion. Note that the sequence begins with PSII. (Yes, you read that correctly. Light is first captured by PSII; the number designations of Photosystems I and II are based on their order of discovery, rather than the order of their roles in PAR capture). The interactive activity Try This! Redox Reactions can only be completed online. The interactive activity Path of Electron and Proton Flow can only be completed online. The interactive activity Study Question 4.2 can only be completed online.

7 Pigments Pigment molecules absorb light energy (photons), to provide energy for the formation of NADPH and ATP. The pigments are contained in the chloroplast light-harvesting complexes (LHC-II and I). Two main classes of pigments are involved in the capture of light energy: chlorophyll and carotenoids. Chlorophyll Chlorophyll a and b absorb blue and red light. Chlorophyll reflects green light, giving plants their green color. These pigments are the main molecules involved in the capture of PAR. Carotenoids Carotenes and xanthophylls are yellow to orange pigments. They participate as accessory pigments in the absorption of blue light. Some carotenoid molecules absorb photons and donate excited electrons to chlorophyll or relay excited electrons between the photosystems. Carotenoids also have a role in the assembly of light-harvesting complexes and also they protect chlorophyll from photo-oxidative damage by attenuating or quenching excessive excitation energy. Fig. 4.5 Absorption spectra of chlorophylls (A) and carotenoids (B).

8 Fig. 4.6 Chlorophyll (Chl) and accessory pigments (Acc) capture light energy. Accessory pigments transfer energy to chlorophyll. Photon absorption converts the pigment from its lowest-energy or ground state to an excited state, causing one of the pigment's electrons to shift its molecular orbit. (Recall that electrons rotate or orbit around the atom's nucleus.) The excited electron can return to its more stable ground state via several mechanisms (Buchanan et al., 2000; Hillier and Babcock, 2001; Ort, 2001). Each mechanism releases the energy in a different form as the electron returns to ground state. Photochemistry The electron from the excited molecule is lost to an electron acceptor molecule, reducing that acceptor molecule. This mechanism transduces light energy into chemical products. Thus, excitation of the pigment initiates the photophosphyration process. Fig. 4.7 Photochemistry of pigment photon absorption and energy transfer. Relaxation Excitation energy is released simply as heat during a non-radiative decay. Carotenoids are involved in thermal energy dissipation (Ort, 2001). Fluorescence Energy is released as light; the light is emitted in a wavelength slightly longer (lower energy) than that of the absorbed light. Energy Transfer This mechanism transfer the energy from one molecule to another, such as from one pigment molecule to another. This is an important and efficient means of energy transfer between antenna and reaction centers. Other names for this mechanism are 'inductive resonance' and 'radiationless transfer'.

9 Fig. 4.8 Diagram of chlorophyll excitation and energy dissipation. Some energy is also lost as heat. What happens when there is excessive light or more energy than the plant can use? Ordinarily, energy is dissipated by combinations of photochemistry, relaxation, fluorescence, and energy transfer mechanisms. Some plants are also able to reduce the amount of light incident on the leaf by altering leaf display and/or chloroplast movement, and thus, reduce photon absorption. However, when the amount of energy absorbed exceeds the energy that the pigments are able to transfer via the usual mechanisms, the photosynthetic apparatus can be damaged Photosystem II is particularly vulnerable. When light energy is excessive, the ph gradient between the lumen and stroma sides of the thylakoid membrane increases (low lumen ph). This change in ph gradient signals a state of excessive energy and may induce a conformational change in carotenoids within the PSII antenna (Ort, 2001). The conformation change, in turn, may promote thermal dissipation of energy, quenching the excessive energy and protecting chlorophyll in the light-harvesting complex (LHC-II). When chlorophyll is in an excessive excitation state, that excited state can last longer and generate damaging molecules, such as excited oxygen molecules and free radicals. These reactive molecules can damage thylakoid membranes, chlorophyll, enzymes, and other molecules, disrupting or interfering with photosynthetic processes. Carotenoids can accept energy from excited chlorophyll and thus, help prevent the formation of oxygen free radicals. Peroxidases and repair mechanisms also enable the plant to rid itself of reactive molecules and repair photo-induced damage. Herbicides are commonly designed to disrupt the light reactions, resulting in eventual plant death. Some herbicides target the biosynthetic pathways involved in the production of carotenoids. Without sufficient carotenoid molecule to provide photo-protection, thylakoid membranes and photosynthetic apparata are damaged. Some herbicides block the electron transport chain of the Z-scheme, interrupting energy transfer. Others autooxidize, forming reactive oxygen species, or free radicals which disrupt membranes. Still other herbicides inhibit photosynthesis by binding to a protein in the thylakoid membrane, stopping ATP and NADPH production and CO 2 fixation.

10 Carbon Dioxide Diffusion Leaf Anatomy Carbon dioxide (CO 2 ) diffuses from the atmosphere into the leaf and to the site of assimilation within the leaf. Re-familiarize yourself with the anatomy of a leaf. In the following activity, click on the terms to view their definitions. Then, click on the boxed area in the graphic for additional terms and definitions. The interactive activity Soybean Leaf Anatomy can only be completed online.

11 The Leaf Diffusion Pathway Gas movement through a leaf presents a unique analytical problem. The pathway involves diffusion through different substances (e.g., air, cell sap), passage through pores (stomata) of varying size, and in the case of CO 2, absorption may be enzyme assisted. Crop physiologists have dealt with this complex circumstance by assuming gas movement is analogous to the flow of electricity, described mathematically by Ohm's Law. Ohm's Law states that flux (gas flow) is directly proportional to the potential gradient (concentration in the case of gas diffusion) and inversely proportional to resistance of the path. In the gas diffusion analogy, flow can be characterized by a series of resistances through the air, stomatal pores and cell sap. In the particular case of CO 2, diffusion for photosynthesis, the simplest formula describing flux is: Equation 4.2 A, assimilation, is the net flux or flow rate of CO 2 into the leaf as measured during photosynthesis; usually A is expressed on a leaf surface (one side) basis in units of µmol CO 2 m -2 s -1. C a refers to the CO 2 concentration of ambient air air that is external to the leaf. C str represents that CO 2 in the chloroplast stroma. The CO 2 transport resistances are partitioned into stomatal resistance, r s, which is determined largely by the density and porosity of stomata; and cellular resistance, r c (sometimes called mesophyll resistance), the resistance internal to the mesophyll cell. A third resistance, boundary layer resistance, r a, is related to diffusion in air external to the leaf; because it is inseparable from rs by conventional measurement techniques and, in most cases, r a is many times smaller than rs, so ra is ignored. Some find it more straightforward to think of the movement of CO 2 in terms of conductance rather than one of resistance. Fig. 4.9 The interactive activity Study Question 4.3 can only be completed online.

12 Measuring the Photosynthetics Characteristics of a Leaf Modern instrument systems use infrared gas analyzers (IRGA) to measure photosynthetic rates because CO2 absorbs infrared light very strongly. The IRGA continuously samples the air flowing from an enclosed, transparent leaf chamber, and compares it to a 'reference sample' taken from the same air stream just before it enters the leaf chamber. Using differential infrared (IR) light absorption, the instrument 'senses' the decline in CO 2 and build-up of water vapor across the chamber by comparing chamber air with reference air. Given these changes of gas concentrations, plus measurements of the leaf and air temperatures, and a value entered by the operator for the leaf surface area, the instrument calculates and displays A; transpiration (Tr); C a ; the leaf internal air CO 2 concentration (C i ); and r s. Fig Schematic representation of typical infrared gas analysis system used for measurement of gas exchange characteristics of leaves. Equation 4.3 This equation requires a value for Cstr to calculate A. Equation 4.2 requires a value for C str to calculate A. This value cannot be measured and must be estimated. A common estimate of C str is approximately 0.5 C a. But this relationship will vary with photosynthetic rate, stomatal resistance, and photosynthetic type. Cellular resistance to CO 2 diffusion is the most difficult to characterize and essentially impossible to measure directly. So it usually is calculated as the residual resistance by subtracting rs from total resistance obtained from Equation 4.2. The stomatal resistance value given by the instrument is calculated for water vapor flux. The rate of water vapor loss (Transpiration, Tr) is estimated by the instrument from a formula similar to that for CO 2 flux. The flux equation for water vapor (Equation 4.4) differs from that for photosynthesis in only one respect there is presumed to be no cellular resistance to water movement. Why? Because the cell surfaces bordering intercellular leaf space are considered to be saturated with water, so that there is no component of cellular resistance, only stomatal resistance in water vapor diffusion.

13 Equation 4.4 Where, Tr is the transpiration rate; W i is the internal concentration of water vapor; W a is atmospheric humidity; and r s H20 is the stomatal resistance to water vapor diffusion. Stomatal resistance for CO 2 flux, r s H20, differs, in absolute value, from that for water vapor flux by the ratio of diffusion coefficients in air for the two gases, 1.6, which is a physical property of the molecules. Equation 4.5 By subtraction, then, we arrive at cellular resistance to CO 2 diffusion, r c. Equation 4.6 Cellular resistance, as estimated by subtraction, probably should more properly be called residual resistance, and should not be considered a true transport resistance because carboxylation (CO 2 fixation) may be the major component of cellular resistance. In plotting A vs. C i, the slope of the linear portion of the relationship is the carboxylation efficiency, 1/r c. Fig Assimilation, A, as a function of internal CO2concentration, Ci, and carboxylation efficiency, 1/rc, the linear portion of A-Ci curve.

14 Carbon Dioxide Assimilation and Sugar Production Carbon dioxide assimilation occurs by three different but related mechanisms in green plants. C3 photosynthesis is the most common among crop plants. The two other mechanisms, C4 photosynthesis and Crassulacean Acid Metabolism (CAM), represent ecological adaptations that improve photosynthetic productivity under stressful conditions. This diversity allows for breadth of ecological adaptation and also is responsible for some, though not all, of the species differences in crop productivity. Study Tip: Be able to describe and explain the similarities and differences among C3, C4, and CAM photosynthetic pathways. Know the main steps and key compound and enzyme names. The biochemistry of these pathways is presented to assist you in understanding the processes involved. However, you are not expected to memorize chemical structures. C3 Photosynthesis C3 photosynthesis gets its name from the first product of CO 2 fixation a three-carbon (3-C) compound, 3-phosphoglycerate or PGA. CO 2 is assimilated in the Photosynthetic Carbon Reduction (PCR) or Calvin Cycle. The PCR cycle has thr ee phases: 1. Carboxylation CO 2 is fixed into a five carbon sugar (5-C), ribulose-1,5-bisphosphate (RuBP) by the enzyme ribulose-1,5-bisphosphate carboxylase-oxygenase, (Rubisco) giving two PGAs; Carboxylation Phase CO 2 is assimilated in the carboxylation phase of the PCR cycle. In the first step, one CO 2 molecule is added to RuBP via a carboxylation reaction. CO 2 is fixed into RuBP by Rubisco forming a six carbon (6-C) intermediate, which is unstable and immediately hydrolyzes to give two molecules of PGA. Phosphate groups, PO32-, are shown as. Each C atom is numbered to help you track their fates. Note that the final step of these reactions is the enzymatic break down of the six carbon intermediate formed by the carboxylation RUBP into two(2) three carbon compounds. These carbon compounds are called phosphoglycerate. Or, 3-phosphoglycerate after remembering the carbons with respect to their 3 carbon structure.

15 3-phosphoglycerate after remembering the carbons with respect to their 3 carbon structure. 2. Reduction PGA is reduced using NADPH and an ATP to another three carbon (3-C) compound, glyceraldehyde 3-phosphate (GAP), a sugar phosphate molecule used for biosynthesis and energy and in the regeneration phase of the PCR cycle. Reduction Phase Each of the 2PGAs are then reduced to make two glyceraldehyde-3-phosphates (GAP) because they contain 3 carbons and a phosphorous group, they are called triose phosphate (TP). So, the carboxylation and reduction reactions yield six carbohydrate carbon atoms, the two TPs, in place of the initial five carbons of RuBP. NOTE however that only one "new" carbon has been added. Both PGA molecules produced by the RuBP carboxylase reaction are reduced to form a molecule of glyceraldehyde 3-phosphate (GAP). Thus, two ATP and two NADPH2 per CO2 molecule fixed are used in the reduction reaction. Note, the reduction reaction for only one of the PGA molecules is shown here. The GAP molecules form PART of the chloroplast TP pool. These TP can be used several different ways depending on the plant/cell carbon balance. Three common uses for the TPs are: 1. to regenerate RUBP to keep the carboxylation/reduction reactions going. 2. TPs can be combined to feed into the hexose (C carbon) phosphate pool. This is necessary in order to support starch synthesis. 3. TPs, perhaps after conversion to dihydroxyacetone phosphate (DHAP), can also be exported from the chloroplast to the cytoplasm via a TP/Pi antiporter. You should have deduced by now that it takes 3 carboxylation/reduction reactions to add 3 "new" carbons to the triose pool. 3. Regeneration RuBP is regenerated through a series of enzyme-mediated reactions, using GAP molecules, water, and ATP. Carboxylation itself does not require input of energy, but the reduction and regeneration phases do. The latter two phases are driven by the chemical energy (NADPH, ATP) generated by the light reactions.

16 Each turn of the PCR cycle: 1. Fixes one CO 2 ; 2. Generates one-sixth of a hexose (6-C) sugar molecule; 3. Regenerates one RuBP molecule (containing five C atoms); and 4. Consumes four hydrogen (H + ) atoms. Fig The PCR cycle has three phases: carboxylation, reduction, and regeneration. (Adapted from Buchanan, 2000). Three CO 2 molecules need to be fixed to yield one "new" GAP molecule. A total of 6 GPA molecules are involved in these 3 cycles. Five GAP molecules are used to regenerate RuBP (recall that RuBP is a 5-C molecule), plus one GAP does into the triose phosphate pool in the stroma. GAP can move into the hexase phosphate and or be converted into dihydroxyacetone phosphate (DHAP) and exported to the cell cytoplasm for use in hexose formation. So again, the assimilation of three molecules of CO 2 produces a net synthesis of a single three-carbon sugar phosphate molecule, GAP. The interactive activity Study Question 4.5 can only be completed online. In summary, the PCR cycle occurs in the stroma of the chloroplast, deriving energy from the light reactions photophosphorylation to assimilate CO 2 into carbohydrates (sugar).

17 Fig The Photosynthetic Carbon Reduction (PCR) cycle of CO2 assimilation. Keep in mind the three phases of the PCR cycle: carboxylation, reduction, and regeneration. The interactive activity Try This! PCR Cycle - Animation can only be completed online. What is the fate of the TP generated in the PCR cycle? Sucrose Synthesis Most of the TP is shuttled from the chloroplast into the cell cytoplasm via the TP-Pi transporter located in the chloroplast membrane. In the cytoplasm, TP is assembled into sucrose, a 12-C sugar, which is loaded into the leaf phloem and exported to growing tissues (Lesson 12). However, not all of the TP exits the chloroplast. RuBP Regeneration As you know, some of the TP must be used to regenerate the CO 2 acceptor molecule, RuBP. Starch Synthesis Some of the TP is also used to produce starch. During the daytime, chloroplasts continually produce starch. During the night, the starch is hydrolyzed back to TP, exported from the chloroplast into the cytoplasm, and assembled into sucrose for export from the leaf to support nighttime plant growth. Thus, another major function of the chloroplast is to synthesize and store starch. The interactive activity Study Question 4.6 can only be completed online.

18 Photorespiration The Process Remember that the name of the enzyme that catalyzes the carboxylation ribulose 1,5 - bisphosphate carboxylase - oxygenase (RUBISCO) is so named because in addition to the carboxylation reaction, the enzyme can also act as an oxygenase reaction. This reaction and those that follow, characterize what is called photorespiration. Photorespiration, refers to light-stimulated evolution of CO 2. Photorespiration is not a true respiration because ATP is not generated as it is in 'dark respiration'. Photorespiration occurs because Rubisco is not specific for CO 2 fixation. About 33% of the time, Rubisco will fix an oxygen molecule instead of CO 2. When this happens, one molecule of a toxic compound, 2-phosphoglycolate (PG), is produced. Fig Oxygenation of RuBP Because the plant cannot utilize PG, it must rid itself of the compound metabolically. The process of breaking down PG involves shuttling compounds between three organelles and ultimately leads to the release of CO 2. The rate of photorespiratory CO 2 loss is estimated between 18 and 27% of the photosynthetic rate. Until recently, photorespiration was believed to have no useful function; but new research suggests that photorespiration may serve several beneficial purposes.

19 Fig PG is formed when Rubisco fixes O2 instead of CO2. PG is shuttled from the chloroplast to the peroxisome to mitochondrion, where the CO2 is released At 25 o C and 350 PPM (Parts Per Million) CO 2, Rubisco will average a ratio of 1 O 2 to 34 CO 2 fixed. The fixation of O 2 obviously does not result in carbon gain, and the PG produced is metabolized through a mechanism that results in additional loss of CO 2 (1 CO 2 for every 2 PG produced). The net consequence of photorespiratory O 2 fixation is a 30% reduction in net carbon gain, with the fixation of O 2 instead of CO 2 accounting for two-thirds of the reduction. The other one-third is a consequence of the loss of CO2 upon decarboxylation of glycine. PPM (Parts Per Million) Parts per million, or ppm, is rarely used now in science writing because it was not included in the international system of scientific units (SI Units). The reason is that ppm has no real meaning unless one specifies whether it is volume/volume, mass/volume, or mass/mass. One needs to use actual units for the meaning to be clear. So, 350 ppm CO 2 in air is more properly written 350 µl CO 2 L -1 air. As temperature increases above 25 o C, photorespiration increases because O2 solubility in cell sap increases faster than does that of CO 2. At temperatures cooler than 25 o C, inhibition by O 2 is

20 proportionately less. But, temperature effects on photorespiration can be masked in whole leaves by direct temperature effects onphotosynthetic enzymes. For example, increasing temperature from 15 o C to 25 o C usually increases net carbon gain in C3 types because Rubisco cycling is accelerated relative to photorespiration. CO 2 - O 2 Competitiveness for Rubisco The CO 2 -O 2 relationship with Rubisco consists of molecules of each gas competing for the same binding site on the enzyme. The enzyme having a much stronger affinity for CO 2 than for O 2. (Oxygen is 600-times more abundant in the atmosphere than is CO 2, yet on average is fixed only 30% of the time.) Since the beginning of the industrial revolution, the CO 2 concentration of the atmosphere has increased about 25%, and with continued burning of fossil fuels at an accelerating rate, it continues to rise rapidly. Fig Graphic representation of the result of CO2 and O2competition for the active site on Rubisco in C3 photosynthesis. G, which is the CO2 compensation atmospheric concentration, is a function of O2 concentration. The red Xmarks the G for each given O2 percentage. In the normal atmosphere of 21% O 2, the CO 2 compensation concentration, G, is about 50 to 80 µl of CO 2 L -1 of air. This is the concentration of CO 2 at which CO 2 fixation by photosynthesis balances the evolution of CO 2 resulting from photorespiration and respiration. Below a concentration of 2%, oxygen is not fixed, there is no photorespiration, and G is zero or nearly so. As oxygen increases above 21%, G increases and photosynthesis declines. Normal atmospheric concentration of CO 2 is about 350 µl L -1 air (350 ppm).

21 C4 Photosynthesis C4 Leaf Anatomy C4 photosynthesis is so named because the first product of CO 2 fixation is a four-carbon (4-C) acid,oxaloacetic acid (OAA). C4 photosynthesis was discovered in sugarcane in the mid-1960's, about a decade and a half after Calvin elucidated the PCR Cycle. Not long afterward, it was noted that photorespiration was not detectable in C4 plants. Some C4 plants feature a unique leaf anatomy, called Kranz anatomy, which facilitates and spatial separation of biochemical functions in photosynthesis. Kranz means "wreath-like" in German; in C4 plants a sheath of large cells surround the vascular bundles, giving a wreath-like appearance in cross-section (Fig. 4.17). Although not all plants exhibit the Kranz anatomy, it helps to illustrate the CO 2 concentrating mechanism and thus we will base our discussion of C4 photosynthesis around it. Fig Comparative leaf anatomy of C3 (top) and C4 (bottom) plants Bundle sheath cells are dense in chloroplasts, contain a lot of starch, and have numerous plasmodesmatal connections with adjacent mesophyll cells. Where a bundle sheath exists in C3 plants, it usually is much reduced in volume and has few or no chloroplasts. C4 Carbon Assimilation Cycle In C4 plants, the PCR Cycle occurs in the bundle sheath cells, but the initial fixation of carbon occurs in mesophyll cells. In mesophyll cells, HCO 3 -, bicarbonate, in the cytoplasm is incorporated into a C

22 compound,phosphoenolpyruvate (PEP), by the enzyme phosphoenolpyruvate carboxylase (PEPCase) to form a 4-C acid, oxalacetic acid (OAA) hence the name C4 photosynthesis. C4 Equilibrium HCO3-. Remember, in liquids CO2 exists in equilibrium with bicarbonate. Fig Initial CO 2 fixation reaction in C4 photosynthesis. Fig Schematic showing locations of C4 photosynthetic reactions. PEPCase fixation of CO 2 occurs in mesophyll cells, and the PCR cycle occurs in bundle sheath cells. The interactive activity Try This! C4 Metabolism can only be completed online. Mesophyll chloroplasts perform the first energy-requiring step, the conversion of OAA to malate, and they also use energy to regenerate the CO 2 acceptor, PEP. Bundle sheath chloroplasts perform PCR

23 Cycle metabolism, including fixation of CO 2 released upon malate decarboxylation and reduction of PGA to triose phosphate (TP). The interactive activity Study Question 4.7 can only be completed online. C4 Plants Avoid CO 2 Loss by Photorespiration. By combining more efficient system for CO 2 fixation with a unique anatomical structure, C4 plants avoid the fixation of O 2 by Rubisco. C4 metabolism essentially pumps CO 2 into the small volume of the bundle sheath, raising the CO 2 concentration around Rubisco many times higher than in the palisade mesophyll cells of a C3 plant. This process essentially denies O 2 access to Rubisco. Although the enzymes for photorespiration are present in C4 plants, photorespiratory CO 2 loss does not occur in significant amounts because of the high concentration of CO 2 in the vicinity of the active site of Rubisco, and the potential to re-fix photorespired CO 2 in the surrounding bundle sheath cells. Thus, C4s generally have higher photosynthetic rates than C3s. As a consequence, C4 plants are more photosynthetically efficient at ambient CO 2 concentrations and high light intensities. Photosynthesis and Plant Adaptation C3 plants are marginally adapted to hot climates that often experience water deficit. Coping with heat and occasional water stresses, plus the 30% loss in carbon acquisition due to photorespiration, threatens their existence in these climates. Some ecologists theorize that C4 plants evolved as an adaptation to stressful climates. C4 plants have better water-use efficiency and use nitrogen more efficiently in photosynthesis than C3s do. Water-Use Efficiency Water-use efficiency is calculated as dry matter gain per unit water used. The greater wateruse efficiency of C4s is not due to their using less water per se. Because C4s do not photorespire they have greater dry matter gains per unit of water used than do C3s. Hot environments tend to be nitrogen poor. Substantial quantities of nitrogen are tied-up in photorespiratory cycling in C3 plants. Lacking this, C4 plants are much more efficient in the use of nitrogen for photosynthesis.

24 Photorespiratory cycling C3-C4 Intermediates For the most part, crop plants are either C3 or C4. In a few genera, both C3 and C4 species occur. One of these, Panicum, is a genus of tropical to temperate grasses. Not only are there distinct C3 and C4 Panicumspecies, there also is a species in which the leaves exhibit both C3 and C4 characteristics a C3-C4 intermediate. However, these plants are mostly curiosities; they have little agronomic worth. The interactive activity Study Question 4.8 can only be completed online. Breeding for Improved Photosynthesis This raises the question: Would it be practical to breed a C4 soybean, or rice, or wheat? Lack of occurrence of C4 species within the many genera of crop plants has precluded breeding C4photosynthesis into traditional C3 crops. Some genera, Triticum (wheat), for example, have been searched without success for the occurrence of C4 variants. Mutagenesis to produce a C4 type in soybean (Glycine) and perhaps in other genera, too, has been tried; but again without success. Recently, genes for C4 photosynthesis have been incorporated successfully into rice (Oryza sativa) a C3 species using transgenic approaches. Transformants expressing PEP carboxylase have slightly higher rates of photosynthesis and decreased sensitivity to O 2 inhibition of photosynthesis (a test for photorespiration). Interestingly, the reason for these improvements was a decrease in stomatal resistance, which apparently resulted in higher concentrations of CO 2 at the site of carboxylation in the mesophyll chloroplasts.

25 Discussion Topic 4.1 What is photorespiration? How is it different from metabolic respiration? Outline the pros and cons of photorespiration. It is a good or bad thing for plants? Discuss why you think plant tolerate a Rubisco enzyme that fixes both CO 2 and O 2. Consider the potential benefits/risks of incorporating a C4 photosynthetic pathway into a C3 plant such as rice. Many millions are being spent in pursuit of this goal. Complete this assignment in Canvas. Be sure to check the course calendar for due dates.

26 Crassulacean Acid Metabolism (CAM) Crassulacean Acid Metabolism (CAM) was named for the plant family in which it was discovered, the Crassulaceae, but it also occurs in many other "succulents". CAM plants exhibit a kind of C4 photosynthesis in which fixation into C4 acids and assimilation via the PCR Cycle are separated temporally, rather than spatially as with C4s. CAM plants can open their stomata at night and assimilate CO 2 into C4 acids, mostly malate, via PEPCase. The malate is stored in vacuoles. In daytime, the malate is released from vacuoles, decarboxylated in the cytoplasm, and the CO 2 is re-fixed via Rubisco in mesophyll chloroplasts. Fig CAM photosynthetic metabolism. The interactive activity Try This! CAM Diurnal Pattern can only be completed online. Another unique aspect of CAM photosynthesis is that the PEP needed for accepting HCO 3 - is generated, in species like pineapple, from vacuolar sugars, which are accumulated in daytime. Finally, CAM differs from C4 in yet a third way. In C4 plants, PEPCase is a light-activated enzyme obviously, that is not true for CAM plants. CAM plants are particularly adapted to desert environments. The rapid decarboxylation of malate in daytime raises the intercellular CO 2 to a high level, causing stomata to close, thus conserving water. Some species are obligate CAM, but many, perhaps most, actually are facultative C3-CAM. During transition periods, early morning and late afternoon, when shifting between day and night metabolism, their stomata are open and CO 2 may be assimilated directly via Rubisco.

27 CAM Water Conservation CAM water conservation. The ability to open stomata and assimilate CO 2 at night when transpirational demand is low, and then close their stomata and metabolize the stored-co 2 of malate in daytime when sunlight energy is available but transpirational demand is great, confers high water-use efficiency and adaptivity of CAM plants to arid and semiarid environments. Through this mechanism, CAM plants are able to cope with conditions that C3 and C4 plants would find extremely stressful, if not intolerable. Crops that utilize CAM generally are less productive than C3 types, but their unique photosynthetic system adapts them to regions where C3 types would be unproductive, even if they could exist. Though CAM crops are not among the food or feed staples that are valued for energy and protein, many are highly valued as ornamentals or for dietary variation. Discussion Topic 4.2 Give your location and list three crops commonly produced in your region. Indicate whether each utilizes C3, C4, or CAM photosynthetic metabolism. Complete this assignment in Canvas. Be sure to check the course calendar for due dates.

28 Summary Understanding the processes involved in photosynthesis is fundamental to understanding crop production and management. Photosynthesis can be separated into three processes: 1. The light reactions solar energy is captured and converted to chemical energy, 2. A diffusion process carbon dioxide moves from the atmosphere through the leaf and into the chloroplast; and 3. A two-step assimilation process carbon dioxide in the chloroplast is fixed and the resulting compound is reduced, using the chemical energy from captured sunlight, to form carbohydrate. The interactive activity Study Question 4.9 can only be completed online. Lesson 4 Reflection 1. In your own words, write a short summary (< 150 words) for this lesson. 2. What is the most valuable concept you learned from the lesson? Why is this concept valuable to you? 3. What concepts in the lesson are still unclear/the least clear to you? Complete this assignment in Canvas. Be sure to check the course calendar for due dates.

29 References Buchanan, B. B., W. Gruissem, R. L. Jones Biochemistry & Molecular Biology of Plants. American Society of Plant Physiologists, Rockville, MD. Hillier, W., and G. T. Babcock Photosynthetic reaction centers. Plant Physiol. 125: Ort, D. R When there is too much light. Plant Physiol. 125:29-32.