The perplexity of life arises from there being too many interesting things in it for us to be interested properly in any of them.
G. K. Chesterton (1874-1936), British author. Tremendous Trifles, "The Secret of a Train" (1909).
Introduction to the concepts of energy and entropy
Who can tell me what "entropy" is? It is the net disorder in any arbitrary system. So far as we know, the universe is becoming increasingly disordered; its entropy is increasing. This has lots of interesting ramifications, which I will not go into here. The one important thing to remember is to reverse increasing entropy requires the expenditure of energy. Thus, all organisms, to live, require sources of energy to do work. Some creationsists say that life violates this law of increasing entropy, thus requiring God's intervention. Discuss the fundamental error of this view.
Introduction to The Sun's energy
Within the sun or any star, a series of nuclear reactions take place where mass is converted to energy in accordance with Einstein's famous equation E = mc2. As a result of these reactions, the sun maintains an extremely high surface temperature (about 6000 degrees absolute); with its huge surface area (8 million miles in diameter) it thus radiates an almost unimaginably large amount of energy into space. Of this radiation, only a tiny amount is incident on the earth (4 ten-billionths) of which only a small amount is absorbed by plants. The energy entering a plant may be lost again or stored by the plant. Of the light absorbed by plant, a fraction is captured as chemical energy. In the context of life on earth, this capture of solar radiation is largely responsible for the functioning and maintenance of the biosphere. The process I am referring to is, of course, photosynthesis, the conversion of light energy into chemical energy and the formation of carbohydrates in plants. Photosynthesis is actually a group of related processes.
As I am sure you already know, light has a wave nature and a particle nature. So if we look at the characteristics of the sun's radiation reaching the earth's surface. First of all, the amount of radiation that actually reaches the outer atmosphere is called the solar constant and is about 2 cal/cm2/min. On a clear day at noon during the summer the amount reaching the earth's surface at this latitude is about 1.3 cal/cm2/min. And if we look at that the composition of incident radiation in terms of wavelength:
99% 150 to 4000 nm are the wavelengths of which
2% UV = 290-380 nm
53% IR = 750-4000 nm
45% = visible 390 to 760 nM which is very similar to the wavelengths absorbed by plants absorb: 380-710 nm.
Thought: is it a coincidence that we see and plant absorb at approximately the same wavelengths? Discuss.
Photosynthesis: The Light-Dependent Reactions
Cholorphyll is a pigment with two structural units. One is a long tail, which is fat-soluble, and dissolves in the membrane lipid bilayer, anchoring the chlorophyll. The other part is a porphyrin ring which encloses a Magnesium atom. It is this ring with alternate single and double bonds interacting with a metal that captures the solar energy.
The fundamental principle of light absorption, often called the Stark-Einstein Law, is that any molecule can absorb only one photon at a time and this photon causes the excitation of one electron. Electrons in stable ground state orbitals are the ones usually excited. An electron in the ground state is in its lowest energy state, that is, it occupies the orbital space of lowest energy. So once excited by a photon, it is driven away from this condition, its ground state, away from the positively charged nucleus a distance corresponding to the energy of the photon that it absorbed. The pigment molecule is then in an excited state. and it is this excitation energy which is going to be captured and stored in photosynthesis. Chlorophylls and other pigments can remain in an excited state only for extremely short periods (on the order of a nanosecond or even less). To simplify what happens to the excitation energy: It can be 1) lost as heat; 2) lost as heat and fluorescence (light production accompanying the rapid decay of excited electrons returning to their ground state - chlorophyll fluorescence produces only deep red light and these long wavelengths can be easily seen when a concentrated solution of either chlorophyll a or chlorophyll b or a mixture is illuminated especially with blue or UV light - fluorescence in leaves is greatly minimized because of the excitation energy being used in PS but can be detected using a fluorometer.
Although the photons are more energetic, blue light is actually harvested less efficiently than red light. After excitation by a blue photon, the electron always decays extremely rapidly by heat release to the lower energy level that red light produces without heat loss when it is absorbed. From this lower level, either additional heat loss, fluorescence, or energy transfer can occur.
An absorption spectrum for a particular pigment describes the wavelengths at which it can absorb light and enter into an excited state.(page 58 in text). An action spectrum, on the other hand, describes the efficiency of a particular photosynthetic system at various wavelengths. This helps to identify which pigments are involved in photosynthesis because these spectra often closely resemble the absorption spectrum of one or more of the participating pigments. When we look at the action spectra for a number of species, we get similar results. All plants show a major peak in the red region and a distinct lower peak in the blue. Both peaks result from light absorption by the chlorophylls. However, many conifers have different action spectra because their waxy needles reflect in the blue region (their needles appear blue-green in color).
Photosynthesis requires that energy in excited electrons of various pigments be transferred to an energy-collecting pigment, the reaction center. There are two reaction centers in plants located in the thylakoids both of which consist of Chlorophyll a molecules that are made special by their association with particular proteins and other membrane components. The energy in an excited pigment can be transferred to an adjacent pigment, and from it to another pigment, and so on until the energy finally arrives at the reaction center. There are various theories which explain the energy migration within a group of neighboring pigments, one of which is called inductive resonance. I won't attempt to explain this theory but I want to emphasize that the excitation of any one of several different pigment molecules in a thylakoid allows momentary collection of the light energy in a Chlorophyll a reaction center.
The other pigments involved in absorbing light are chlorophyll b and c, the carotenoids (carotenes and xanthophylls) and the phycobillins. These are known as accessory pigments, and the energy they absorb must be transferred to Chlorophyll a to be captured permanently.
How does chlorophyll transfer its excited state into useful biosynthetic energy?
A system of membrane-bound enzymes transfers the electrons taken by chlorophyll from water down an energy pathway, pumping hydrogen ions across the thylakoid membrane. This generates a potential energy difference (proton gradient) across the thylakoid membrane, due to the concentration of positive charge on the side to which the hydrogen has been transferred. This charge separation is alleviated by allowing the hydrogen ions to pass back through the membrane through an enzyme pump that generates ATP with the electric potential produced by the charge separation. The electrons are finally transferred to NADP, producing NADPH which can be used as a source of reductive power for biosyntheses.
Non-Cyclic Photophosphorylation
The form of photosynthesis with which we are most familiar is non-cyclic Photophosphorylation (figure 10-14 and 10-17). It consists of two sets of thylakoid-embedded pigments to excite. They are called PS1, or photosystem 1, and PS2, or photosystem 2. PS1 is best excited by light at 700 nm, and is thus sometimes called P700. PS2 is best excited by light at 680 nm, and is thus sometimes called P680. The Z-shape is important because it is an indicator of the relative energy state of the electrons during this process. Anytime they go downhill, they are releasing energy; anytime the arrows go up, they are being boosted to a higher energy level by absorbed light.
Energy enters the system when PS2 becomes excited by light. It grabs a pair of electrons from water, producing a molecule of oxygen gas for every two waters split. These "boosted" electrons are shed by the excited PS2, returning it to its unexcited state. The electrons are passed "downhill" through a chain of oxidation-reduction reactions, called an electron transport chain. The electron transport components of the model can be compared to the bucket brigade. Just as people in a bucket brigade rapidly move buckets of water toward a fire, so these components move electrons rapidly from H2O to NADP+.
Each element in the pathway is reduced by the electron, and turns right around to reduce its neighbor in the pathway by giving it the electron, thus becoming reoxidized and ready for the next electron to pass through the photosystem. At each step, the electron moves to a lower energy level.
What is actually happening in the thylakoid membrane? Look at Figure 10-16 from left to right. It is a diagram of the "chemiosmotic coupling mechanism" of photophosphorylation. It is PS2 that provides the energy boost to move the electron through the first electron transport chain, thus pumping protons through the membrane into the lumen. At the far right, you cansee the ATP synthetase molecule in the membrane, which allows protons back trough the membrane and harvests this energy to generate ATP.
PS1, on the other hand, passes on to a different energy transport chain the energy required to produce NADPH. This division of labor between the two photosystems becomes important when we look at cyclic photophosphorylation.
To summarize, electrons move in the following way:
PS2 -----> PS1 ------> NADPH + H (H2O is oxidized from light energy derived from PS2, and two electrons are cooperatively transferred to PS1 and on to NADPH.).
Besides these two photosystems two other major constituents of the thylakoid membranes are the light harvesting complexes (LHC I and LHC II). They are composed of Chlorophyll a and b, xanthophyll and carotenoids and their function is to harvest light energy by absorbing it and transferring it to the proper photosystem. Another function of accessort pigment is thought to be protection of the active pigments from light damage.
Looking at thylakoids, they form grana (each granum being a disk-like stack of thylakoids). The thylakoids that do not form stacks are called stroma thylakoids. In general PS2 units are located within the stacks of grana and PS1 are found within the stroma thylakoids.
Another important component of thylakoids necessary for photophosphorylation (the light driven formation of ATP) is a complex of proteins called the coupling factor. This complex synthesizes ATP from ADP and P and the process is driven by light driven electron transport. For each ATP formed 3 protons are transported from the lumen back to the stroma.
Cyclic Phosphorylation
Sometimes an organism is short of energy, and in a situation where it is too starved to think about reductive power to synthesize new molecules and to grow. All it wants is energy to use to fulfill basic functions. Many bacteria can shut off PS2, and in fact some do not even have it. This allows them to produce ATP in the absence of glucose by generating a proton gradient across the membrane using the mechanisms of photosynthesis. This type of energy generation is called cyclic photophosphorylation.
Cyclic electron flow is the process where electrons pass through PSI and once they have reached and reduced ferredoxin, instead of passing downhill to NADP, cycle back to the plastoquinones of PSII.. It is thought to occur when the cell is believed to have enough reducing power, that is NADPH, but requires additional ATP for other metabolic needs. In cyclic electron flow, no water is split and thus no O2 is evolved, but ATP is still produced.
This may seem counter-intuitive. It appeared from noncyclic photophosphorylation that PS1 was responsible for NADPH production, while in cyclic photophosphorylation it is important for ATP production. This apparent dichotomy can be resolved when we understand what makes PS1 both a good candidate for noncyclic photophosphorylation and for NADPH production. PS1 is very good at transferring an electron, whether it be to NADP or to ferredoxin (fd). It is a powerful reductant. PS2, on the other hand, is better at grabbing electrons from water to transfer them to quinone (Q). It is a good oxidant. Thus, in cyclic phosphorylation, the electron transferred is not derived from water, but from PS1 itself. It therefore must be recycled to PS1.
Photosynthesis: The Light-Independent Reactions
In the second stage of photosynthesis, the chemical energy harvested by the light-dependent reactions is used to reduce carbon from CO2 to 6-carbon sugars. There are two major pathways of carbon fixation: the Calvin Cycle (or C3 pathway) and the C4 pathway. C3 plants use only the Calvin cycle, while C4 plants use both cycles.
The Calvin Cycle
In the C3 pathway, CO2 or O2 is fixed in the leaf by the enzyme Ribulose 1,5-bisphosphate carboxylase/oxygenase, called "Rubisco" for short. See diagram 10-18 on page 281.
1. Ribulose 1,5-bisphosphate (RuBP a 5-carbon sugar with 2 phosphate groups) reacts with CO2 to form a 6-carbon intermediate which is immediately hydrolyzed to 2 molecules of 3-phosphoglycerate (called PGA). Rubisco catalyzes both the attachment of the CO2 to RuBP and the simultaneous hydrolysis of the 6-carbon sugar.
2. Rubisco also catalyzes the oxygenation of RuBP in which O2 reacts with RuBP to form only one 3-PGA and one phosphoglycolate -----> photorespiration. So O2 competes directly with CO2 for the substrate RuBP and at the same site for catalysis on Rubisco. This reaction limits the efficiency of photosynthesis, since only one molecule of PGA is formed from RuBP rather than 2. Under conditions of water stress, when stomata are closed, CO2 is used up and oxygen accumulates, photorespiration reduces photosynthetic efficiency.
3. The cycle runs 6 times, each time incorporating a new carbon and regenerating RuBP. Ribulose is a five-carbon sugar and the gylceraldehydes are three-carbon sugars. Running through the cycle six times generates:
6(5-carbon sugars) + 6(incorporated carbon dioxides)
Those six carbon dioxides are reduced to glucose by the conversion of NADPH to NADP+. Glucose can now serve as a building block to make polysaccharides, other monosaccharides, fats, amino acids, nucleotides, and all the molecules living things require.
Rubisco Described: It is the most abundant protein in the universe, okay on earth. Can make 25-50% of the total N. It is a soluble protein found in the stroma of chloroplasts at very high concentrations. It is inefficient. This comes from the observation that if CO2 is increased in the atmosphere surrounding a plant which is also under high light there is an increase in CO2 uptake in most plants. This indicates that CO2 availability and its incorporation by Rubisco are the limiting step in carbon fixation and not the light reactions. So at air levels of CO2 (0.035%) and O2 (21%) two to three moles of CO2 are fixed for every one mole of O2.
The C4 pathway
The inefficiency above suggests that one way to increase photosynthetic efficiency would be to raise the CO2 conc. around the enzyme. And this is exactly what some plants have been able to do by a rather an elaborate strategy. These plants have been called C4 plants because of the CO2 is first fixed not by Rubisco but by another enzyme, PEP carboxylase and the product formed is a four carbon compound. Most plants which initially fix Rubisco are called C3 plants because the initial products form 3 carbon molecules. C4 plants can virtually eliminate photorespiration by their special biochemistry and leaf anatomy.
Leaf anatomy: , 1882, a German Habberlandt, was first to notice the distinguishing leaf anatomy of C4 plants. If you look at a cross-section of a C4 leaf, the bundle sheath cells which surround the vascular bundle are distinctive because in contrast to C3 plants they contain a large number of organelles (especially chloroplasts and mitochondria). These bundle sheath cells (BSC) also possess very thick secondary cell walls which in some plants can contain a suberin. He suggested that there may be a division of labor between these BSC and mesophyll cells.
Biochemistry: Hatch and Slack, 1960's. What C4 plants have managed to do is to spatially isolate the enzyme Rubisco from the atmosphere. Rubisco is located only within the stroma of the chloroplasts of the bundle sheath cells. So in chloroplasts of the mesophyll cells in C4 plants, there is no Rubisco. But Rubisco is still necessary for carboxylation of CO2.
How does the plant deliver CO2 to this enzyme to Rubisco which is now virtually blocked off from the surrounding environment? The CO2 pump is the reaction of HCO3 in the cytoplasm of mesophyll cells where CO2 reacts with PEP to form OAA an Pi. OAA is quickly converted to malate or transaminated with alanine to aspartate and pyruvate. Bottom line: in C4 plants, there is a division of labor. Mesophyll cells use pepcarboxylase which has a high affinity and Bundle sheath Cells have Rubisco. Asp or mal diffuse into the BSC where they are decarboxylated by specific enzymes depending on the species. Thus CO2 is released in the BSC for the Calvin Cycle.
Ultimately, net photosynthetic rate in C4 plants can be 2-3 times that of C3 plants; they use CO2 more efficiently, and their stomates are consequently smaller. This has the physical consequence that they transpire (use and require)less water. In addition, the optimum temperature range is higher. Thus they are adapted to hotter, drier conditions.
The CAM pathway
Yet a third carbon fixation pathway is found in desert plants: Crassulacean Acid Metabolism (CAM). This mechanism allows plants to fix CO2 as a four-carbon compound during the night and store it temporarily in vacuoles. During the day when stomates are closed, this carbon is released into the cells and fixed normally vias the C3 pathway. Thus it is similar to C4 plants, but instead of using C3 and C4 pathways is physically isolated locations, CAM plants use the C4 pathway at night, and the C3 pathways during the day.
Respiration
Respiration is the process whereby chemical energy stored in carbohydrates is transferred to ATP, the universal energy-carrying molecule. In that form, energy is available to do work of all sorts in the cell. Respiration involves three main stages, glycolysis, the Krebs Cycle, and the electron transport chain. In glycolysis, the 6-carbon glucose molecule is broken down to a pair of 3-carbon molecules of pyruvate. In the Krebs Cycle, the pyruvates are oxidized to CO2 and water. The resulting electrons are passed to an Electron transport chain, where the electrons cascading down the pathway are used to produce ATP from ADP. It is essentially the reverse of photosynthesis, and takes place not only in plants but in animals as well. There is a chemiosmotic coupling mechanism in respiration similar to that described for photosynthesis, called oxidative phosphorylation.
Respiration can also occur anaerobically, when it is called fermentation.
Some important things to remember:
a) overall equations for photosynthesis and respiration
b) describe non-cyclic electron flow through PS2 and PS1
c) define chemiosmotic coupling and describe the role of ATP-synthase in both respiration and photosynthesis
d) the relationship between wavelength and energy in light
e) the major pigments and their roles