Photosynthesis in Higher Plants

• Green plants carry out ‘photosynthesis’, a physico-chemical process by which they use light
  energy to drive the synthesis of organic compounds.
• Photosynthesis is important due to two reasons:
  i. it is the primary source of all food on earth.
  ii. responsible for the release of oxygen into the atmosphere.

WHAT DO WE KNOW?

• Chlorophyll (green pigment of the leaf), light and CO2 requires for photosynthesis. Experiment for Light :-
• In two leaves – a variegated leaf or a leaf that was partially covered with black paper, and exposed to light. On testing these leaves for the presence of starch it was clear that photosynthesis occurred only in the green parts of the leaves in the presence of light. Experiment for Carbon dioxide:-
• A part of a leaf is enclosed in a test tube containing some KOH soaked cotton (which absorbs CO2), while the other half is exposed to air.
• Parts of the leaf, you must have found that the exposed part of the leaf tested positive for starch while the portion that was in the tube, tested negative. This showed that CO2 required for photosynthesis.
  

EARLY EXPERIMENTS

• Joseph Priestley (1733-1804) in 1770 performed a series of experiments that revealed the essential role of air in the growth of green plants.
• Priestley discovered oxygen in 1774.

• Jan Ingenhousz (1730-1799) showed that sunlight is essential to the plant process that somehow purifies the air fouled by burning candles or breathing animals.
• Ingenhousz in an elegant experiment with an aquatic plant showed that in bright sunlight, small bubbles were formed around the green parts these bubbles to be of oxygen.
• Showed that it is only the green part of the plants that could release oxygen.
• 1854 that Julius von Sachs provided evidence for production of glucose when plants grow.
• Glucose is usually stored as starch.
• He showed that the green substance in plants (chlorophyll as we know it now) is located in special bodies (later called chloroplasts) within plant cells.
• T.W Engelmann (1843 – 1909). Using a prism he split light into its spectral components and then illuminated a green alga, Cladophora, placed in a suspension of aerobic bacteria.
• The bacteria were used to detect the sites of O2 evolution. He observed that the bacteria accumulated mainly in the region of blue and red light of the split spectrum.
• Cornelius van Niel (1897-1985), who, based on his studies of purple and green bacteria, demonstrated that photosynthesis is essentially a light-dependent reaction in which hydrogen
  from a suitable oxidizable compound reduces carbon dioxide to carbohydrates.
• When H2S is the hydrogen donor for purple and green Sulphur bacteria, the ‘oxidation’ product is Sulphur or sulphate depending on the organism but not Oxygen. (NEET 2018)
• The Oxygen released from water; this was proved using radio isotope techniques.

WHERE DOES PHOTOSYNTHESIS TAKE PLACE?

• Photosynthesis does take place in the green leaves of plants but it does so also in other green
  parts of the plants.
• Chloroplasts align themselves along the walls of the mesophyll cells, such that they get the
  optimum quantity of the incident light.
• Within the chloroplast there is membranous system consisting of grana, the stroma lamellae,
  and the matrix stroma. There is a clear division of labour within the chloroplast.
• Light reactions (photochemical reactions) : membrane system is responsible for trapping the light energy and also for the synthesis of ATP and NADPH directly light driven . 177 / 302
• Dark reactions (carbon reactions) : In stroma, enzymatic reactions synthesize sugar, which in
  turn forms starch are not directly light driven but are dependent on the products of light
  reactions (ATP and NADPH).

HOW MANY TYPES OF PIGMENTS ARE INVOLVED IN PHOTOSYNTHESIS?

• A chromatographic separation of the leaf pigments shows that the colour that we see in leaves is not due to a single pigment but due to four pigments: Chlorophyll a (bright or blue green in the chromatogram), chlorophyll b (yellow green), xanthophylls (yellow)
  carotenoids (yellow to yellow-orange).
• the wavelengths at which there is maximum absorption by chlorophyll a, i.e., in the blue and the red regions, also shows higher rate of photosynthesis.
• Chlorophyll a is the chief pigment associated with photosynthesis.
• most of the photosynthesis takes place in the blue and red regions of the spectrum.
• Chlorophyll is the major pigment responsible for trapping light, other thylakoid pigments like chlorophyll b, xanthophylls and carotenoids, which are called accessory pigments, also absorb light and transfer the energy to chlorophyll a.

WHAT IS LIGHT REACTION?

• ‘Photochemical’ phase include light absorption, water splitting, oxygen
  release, and the formation of high-energy chemical intermediates, ATP
  and NADPH. Several protein complexes are involved in the process.
• pigments are organized into two discrete photochemical light harvesting complexes (LHC)
  i. Photosystem I (PS I) ii. Photosystem II (PS II).
• These are named in the sequence of their discovery, and not in the sequence in which they function during the light reaction. 178 / 302
• The LHC are made up of hundreds of pigment molecules bound to proteins.
• Each photosystem has all the pigments (except one molecule of chlorophyll a) forming a light
  harvesting system also called antennae.
• The single chlorophyll a molecule forms the reaction centre.
• In PS I the reaction centre chlorophyll a has an absorption peak at 700 nm, hence is called
  P700.
• In PS II it has absorption maxima at 680 nm, and is called P680.

THE ELECTRON TRANSPORT

• In photosystem II the reaction centre chlorophyll a absorbs 680 nm wavelength of red light
  causing electrons to become excited and jump into an orbit farther from the atomic nucleus.
• Electrons are picked up by an electron acceptor which passes them to an electrons transport system consisting of cytochromes.
• This movement of electrons is downhill, in terms of an oxidation-reduction or redox potential scale.
• Electrons are not used up as they pass through the electron transport chain, but are passed on to the pigments of photosystem PS I.
• Simultaneously, electrons in the reaction centre of PS I are also excited when they receive red light of wavelength 700 nm and are transferred to another accepter molecule that has a greater redox potential.
• These electrons then are moved downhill again, this time to a molecule of energy-rich NADP+.
  . The addition of these electrons reduces NADP+ to NADPH + H+.
• This whole scheme of transfer of electrons, starting from the PS II, uphill to the acceptor, down
  the electron transport chain to PS I, excitation of electrons, reducing it to NADPH + H+ is called
  the Z scheme, due to its characteristic shape.
• This shape is formed when all the carriers are placed in a sequence on a redox potential scale.

Splitting of Water

• Electrons that were moved from photosystem II must be replaced. This is achieved by electrons available due to splitting of water.
• The splitting of water is associated with the PS II; water is split into 2H+, [O] and electrons.
• Creates oxygen, one of the net products of photosynthesis.
• Electrons needed to replace those removed from photosystem I are provided by photosystem II located on the inner side of the membrane of the thylakoid.

Cyclic and Non-cyclic Photo-phosphorylation

Living organisms have the capability of extracting energy from
oxidisable substances and store this in the form of bond energy.

• Phosphorylation - ATP is synthesised by cells (in mitochondria
  and chloroplasts).
• Photophosphorylation - is the synthesis of ATP from ADP and
  inorganic phosphate in the presence of light.
• Non-cyclic photo-phosphorylation - When the two photosystems work in a series, first PS II and then the PS I,
• The two photosystems are connected through an electron
  transport chain, as seen earlier – in the Z scheme. Both ATP and NADPH + H+ are synthesised by this kind of electron flow.
• Cyclic photo-phosphorylation - When only PS I is functional, the electron is circulated within the
  photosystem and the phosphorylation occurs due to cyclic flow of electrons .
• results only in the synthesis of ATP, but not of NADPH + H+. (Aipmt 2009,2010)
• occurs when only light of wavelengths beyond 680 nm are available for excitation.
• possible location where this could be happening is in the stroma lamellae.
• While the membrane or lamellae of the grana have both PS I and PS II the stroma lamellae membranes lack PS II as well as NADP reductase enzyme.
• The excited electron does not pass on to NADP+ but is cycled back to the PS I complex through the electron transport chain

Chemiosmotic Hypothesis

Chemiosmotic hypothesis explain the mechanism actually ATP is synthesized in the chloroplast.

• ATP synthesis is linked to development of a proton gradient across a membrane (membranes
  of thylakoid).
• the proton accumulation is towards the inside of the membrane, i.e., in the lumen.
• the processes that take place during the activation of electrons and their transport to
  determine the steps that cause a proton gradient to develop :-

(a) Since splitting of the water molecule takes place on the inner side of the membrane, the protons or hydrogen ions that are produced by the splitting of water accumulate within the lumen of the thylakoids.

(b) As electrons move through the photosystems, protons are transported across the membrane. This
happens because the primary accepter of electron which is located towards the outer side of the
membrane transfers its electron not to an electron carrier but to an H carrier. Hence, this molecule removes a proton from the stroma while transporting an electron. When this molecule passes on its electron to the electron carrier on the inner side of the membrane, the proton is released into the inner side or the lumen side of the membrane.

(c) The NADP reductase enzyme is located on the stroma side of the membrane. Along with electrons that come from the acceptor of electrons of PS I, protons are necessary for the reduction of NADP+ to NADPH+ H+ .protons are also removed from the stroma.

• Within the chloroplast, protons in the stroma decrease in number, while in the lumen there is
  accumulation of protons. (NEET 2016)
• This creates a proton gradient across the thylakoid membrane as well as a measurable
  decrease in pH in the lumen
• This gradient is important because it is the breakdown of this gradient that leads to the
  synthesis of ATP.
• gradient is broken down due to the movement of protons across the membrane to the stroma
  through.

• The transmembrane channel of the CF0 of two parts:
  i. one called the CF0 of the ATP synthase.
• ATP synthase enzyme consists is embedded in the thylakoid membrane and forms a
  transmembrane channel that carries out facilitated diffusion of protons across the membrane.
  ii. other portion is called CF1 and protrudes on the outer surface of the thylakoid membrane on
  the side that faces the stroma.

Break down of the gradient provides enough energy to cause a conformational change in the
CF1 particle of the ATP synthase, which makes the enzyme synthesise several molecules of
energy packed ATP.

Summary of Chemiosmosis

Chemiosmosis requires a membrane, a proton pump, a proton gradient and ATP synthase.
Energy is used to pump protons across a membrane, to create a gradient or a high
concentration of protons within the thylakoid lumen. ATP synthase has a channel that allows
diffusion of protons back across the membrane; this releases enough energy to activate ATP
synthase enzyme that catalysis the formation of ATP.

WHERE ARE THE ATP AND NADPH USED?

▪ The primary acceptor of carbon dioxide

• the acceptor molecule was a 5-carbon ketose sugar – ribulose bisphosphate (RuBP).

The Calvin Cycle

Calvin and his co-workers then worked out the whole pathway and showed that the pathway
operated in a cyclic manner; the RuBP was regenerated.

• Calvin pathway occurs in all photosynthetic plants; it does not matter whether they have C3 or
  C4 (or any other) pathways.

Calvin cycle can be described under three stages:

(1) Carboxylation

(2) Reduction

(3) Regeneration

(1) Carboxylation :

• Carboxylation is the fixation of CO2 into a stable organic intermediate.

• Carboxylation is the most crucial step of the Calvin cycle where CO2 is utilised for the carboxylation of RuBP.
• This reaction is catalyzed by the enzyme RuBP carboxylase which results in the formation of two
  molecules of 3-PGA.
• enzyme also has an oxygenation activity it would be more correct to call it RuBP carboxylase-oxygenase or RuBisCO.

(2) Reduction :

• a series of reactions that lead to the formation of glucose.

• involve utilisation of 2 molecules of ATP for phosphorylation and two of NADPH for reduction
  per CO2 molecule fixed.
• fixation of six molecules of CO2 and 6 turns of the cycle are required for the formation of one
  molecule of glucose from the pathway.

(3) Regeneration :

• Regeneration of the CO2 acceptor molecule RuBP is crucial if the cycle is to continue uninterrupted.

• Regeneration steps require one ATP for phosphorylation to form RuBP
• Hence for every CO2 molecule entering the Calvin cycle, 3 molecules of ATP and 2 of NADPH
  are required.

The C⁴ Pathway

Plants have the C4 oxaloacetic acid as the first CO2 fixation product they use the C3 pathway
or the Calvin cycle as the main biosynthetic pathway.

• C⁴ plants are special: They have a special type of leaf anatomy, they tolerate higher
  temperatures, they show a response to high light intensities, they lack a process called
  photorespiration and have greater productivity of biomass.
• Particularly large cells around the vascular bundles of the C4 plants are called bundle sheath
  cells, and the leaves which have such anatomy are said to have ‘Kranz’ anatomy. ‘Kranz’ means
  ‘wreath’ and is a reflection of the arrangement of cells.
• bundle sheath cells may form several layers around the vascular bundles; they are characterised by having a large number of chloroplasts, thick walls impervious to gaseous
  exchange and no intercellular spaces. (NEET 2011) Ex. - leaves of C4 plants – maize or sorghum – to observe the Kranz anatomy and the distribution of mesophyll cells.
• Hatch and Slack Pathway, is again a cyclic process (Fig 13.9)
• The primary CO2 acceptor is a 3-carbon molecule Phosphoenol pyruvate (PEP) and is present in the mesophyll cells. (NEET 2017)
• Enzyme responsible for this fixation is PEP carboxylase or PEPcase.
• the mesophyll cells lack RuBisCO enzyme.
• The C4 OAA is formed in the mesophyll cells.
• forms other 4-carbon compounds like malic acid or aspartic acid in the mesophyll cells itself, which are transported to the bundle sheath cells.
• In the bundle sheath cells these C4 acid are broken down to
  184 / 302 release CO2 and a 3-carbon molecule.
• The 3-carbon molecule is transported back to the mesophyll where it is converted to PEP again,
  thus, completing the cycle.
• Bundle sheath cells rich in an enzyme Ribulose bisphosphate carboxylase-oxygenase
  (RuBisCO), but lack PEPcase.
• The basic pathway that results in the formation of the sugars, the Calvin pathway, is common
  to the C3 and C4 plants.

Photorespiration

• The process that creates an important difference between C3 and C4 plants is
  Photorespiration. (NEET 2012,2016)
• RuBisCO that is the most abundant enzyme in the world that its active site can bind to both
  carbon dioxide and oxygen.
• RuBisCO has a much greater affinity for CO2 compare to Oxygen.
• in C3 plants some oxygen does bind to rubisco and hands carbon dioxide fixation is decreased.
• RuBP instead of being converted to two molecule of PGA bind with oxygen to form 1 molecule and phosphoglycolate in a pathway called photorespiration .
• photorespiratory pathway there is no synthesis of ATP or NADPH. The biological function of
  photorespiration is not known yet.
• In C4 plants photorespiration does not occur so Productivity and yield are better in these
  plants. (NEET 2016)

Factors affecting Photosynthesis

• The plant factors include the number, size, age and orientation of leaves, mesophyll cells and
  chloroplasts, internal CO2 concentration and the amount of chlorophyll.
• The plant or internal factors are dependent on the genetic predisposition and the growth of the
  plant.
• External factors would include the availability of sunlight, temperature, carbon dioxide
  concentration and water. (also effect stomata movement) (NEET 2018)
• As a plant photosynthesises, all these factors will simultaneously affect its rate.
• Carbon dioxide major factor that limits the rate of photosynthesis. 185 / 302
• Blackman’s (1905) Law of Limiting Factors
• If a chemical process is affected by more than one factor, then its rate will be determined by
  the factor which is nearest to its minimal value: it is the factor which directly affects the process
  if its quantity is changed.

Light

• linear relationship between incident light and CO2 fixation rates at low light intensities.
• At higher light intensities, gradually the rate does not show further increase as other factors become limiting.
• light saturation occurs at 10 per cent of the full sunlight. (NEET 2017)
• Except for plants in shade or in dense forests, light is rarely a limiting factor in nature.
• Increase in incident light beyond a point causes the breakdown of chlorophyll and a decrease in photosynthesis.

Carbon dioxide concentration

• Carbon dioxide is the major limiting factor for photosynthesis.
• The concentration of CO2 is very low in the atmosphere (between 0.03 0.04 per cent).
• Increase in concentration upto 0.05 per cent can cause an increase in CO2 fixation rate after
  that it start damaging the rate. (NEET 2017)
• The C3 and C4 plants respond differently to CO2 .
• C⁴ plants show saturation at about 360 μlL-1
• C3 responds to increased CO2 concentration and saturation is seen only beyond 450 μlL-1.
• Current availability of carbon dioxide level is limiting to C3 plants.
• increased rates of photosynthesis leading to higher productivity has been used for some greenhouse crops such as tomatoes and bell pepper. (NEET 2017) Ex. of C3 plant tomatoes and bell pepper.

Temperature

The dark reactions being enzymatic are temperature controlled.

• the light reactions are also temperature sensitive they are affected to a much lesser extent.
• The C4 plants respond to higher temperatures and show higher rate of photosynthesis while
  C3 plants have a much lower temperature optimum. (NEET 2016,2017)

Water

• Water stress causes the stomata to close hence reducing the CO2 availability.
• water stress also makes leaves wilt, thus, reducing the surface area of the leaves and their
  metabolic activity as well.

NEET 2017 Questions

Q -1. With reference to factors affecting the rate of photosynthesis, which of the following

statements is not correct?

(a) Light saturation for CO2 fixation occurs at 10% of full sunlight

(b) Increasing atmospheric CO2 concentration upto 0.05% can enhance CO2- fixation rate.

(c) C3 - plants respond to higher temperature with enhanced photosynthesis, while C4 - Plants

have much lower temperature optimum

(d) Tomato is a greenhouse crop, which can be grown in CO2, enriched atmosphere for higher

yield.

NEET 2018 Questions

Q-1. Oxygen is not produced during photosynthesis by

(a) Cycas

(b) Nostoc

(c) Green sulphur bacteria

(d) Chara

Q-2. Which of the following is not a product of light reaction of photosynthesis?

(a) NADPH

(b) NADH

(c) ATP

(d) Oxygen

187 / 302

Q–3. Stomatal movement is not affected by

(a) Oxygen concentration

(b) Light

(c) Temperature

(d) Carbon dioxide concentration