Photosynthesis: B. Sc. II Semester- IV Paper No. VIII

Department of Botany

Class: B. Sc. II   Semester- IV Paper No. VIII

 

Photosynthesis

Introduction:

Photosynthesis is one of the most important biochemical process because the life on the earth depend upon this. It is the only process in which solar/radiant energy is absorbed and converted into chemical energy in the form of food material.

Photosynthesis (Greek word Photon = light and Synthesis = to build or putting together) can be defined as the green plants in presence of sunlight and with the help of simple inorganic substances like CO₂ and H₂O produces complex organic substances as glucose with release of O₂ as byproduct. The overall reaction of photosynthesis is

 

 

 

 

 

Photosynthesis is also known as anabolic process (synthesis of complex substances) and oxido-reduction process or redox process (H₂O gets oxidation and CO₂ reduction). The main importance of photosynthesis is the formation of food material and release of O₂ which are useful in maintaining the life on the earth.

 

Site of photosynthesis:

 

All biochemical reactions of photosynthesis take place in the chloroplasts hence chloroplast is the main site of photosynthesis. The chloroplast is a double membrane bound cell organelle and show outer envelope and inner envelope. In higher plants they are disc like or lens shaped and about 4 – 10 mµ in length and 2 -4 mµ in diameter. It encloses a colourless, colloidal matrix called stroma. The stroma contains 70 s ribosomes, DNA and enzymes for reduction of CO₂ into glucose. DNA is circular, closed, naked ring like known as plastidome. As chloroplast contains its own DNA it is a semi-autonomous cell organelle.

The stroma also shows a distant lamellar system which is formed from a small sac like structure known as thylakoid. At certain regions thylakoids are arranged one above the other like a stack of coins which forms a grana lamellae. The number of grana lamellae per chloroplast varies and approximately range from 40 -60. The photosynthetic pigments are embedded in the grana lamellae. The grana lamellae are interconnected with each other by tubular structure called stroma lamellae or inter-grana or fret membrane. Enzymes of dark reaction are present in stroma lamellae hence dark reaction take place in them.

According to Park and Baggins, the inner surface of thylakoid membrane in the grana region show many small particles. These are known as Quantasomes. Quantasome is a photosynthetic unit and it contains all photosynthetic pigments to carry out the photochemical act.

 

 

Photosynthetic pigments:

Photosynthesis is a photochemical reaction in which solar/radiant energy is absorbed and converted into chemical energy. This process is called photochemical act. The pigments which are involved in photochemical act (photosynthesis) are known as photosynthetic pigments.

According to chemical nature, there are three different types of photosynthetic pigments namely Chlorophylls, Carotenoids and Phycobilins.

1) Chlorophylls: These are green in colour and most important as well as aboundant pigment of photosynthesis. The chlorophylls are insoluble in water but soluble in organic solvents. There are seven different types of chlorophylls named as chl. a, chl. b, chl. c, chl. d, chl. e, bacterio-chlorophyll a and bacterio-chlorophyll b (bacterio-viridin). The distribution of photosynthetic pigments in plant kingdom is as follow-

 

Sr. No.	Type of pigment	Distribution in plant kingdom
1	Chlorophyll- a	All green plants except photosynthetic bacteria
2	Chlorophyll- b	All higher plants, Chlorophyta, Euglenophyta
3	Chlorophyll- c	Chrysophyta, Phaeophyta, Pyrrophyta
4	Chlorophyll- d	Rhodophyta
5	Chlorophyll- e	Xanthophyta
6	Bacteriochlorophyll- a	Purple sulphur bacteria
7	Bacteriochlorophyll- b	Green sulphur bacteria

 

Out of these, chl. a and chl. b are best known and most aboundantly found in all autotrophic organisms. The chlorophylls mainly functions for absorption of light energy and converted into chemical energy.

Each chlorophyll molecule is a porphyrin derivative and shows two parts as head and tail. The head region contains a tetrapyrol rings, containing Magnesium (Mg ²⁺) at the center and at one end it has a long phytol chain of carbon and hydrogen. The molecular formula of chl. a is C₅₅H₇₂O₅N₄Mg and that of chl. b is C₅₅H₇₀O₆N₄Mg.

The main difference between chl. a and chl. b is found at carbon number 3. The chl. a have methyl (CH₃) group while chl. b have aldehyde (CHO) group. The chl. a and chl. b show maximum absorption in blue violet and red region of visible spectrum of light.

 

 

                       

 

 

2) Carotenoids: These are present in variable concentrations in nearly all higher plants, green algae, red algae and photosynthetic bacteria. Carotenoids show wide range of colour from yellow, orange to red. They are insoluble in water but soluble in organic solvents. Carotenoids contains a long chain of hydrocarbon linked by conjugated single or double bonds with six carbon rings at each end. They mainly absorbs blue-violet radiations.

            Carotenoids are also further classified into two types as

a) Carotene: These are hydrocarbons, orange in colour and with empirical formula as C₄₀H₅₆. The name of carotene ends with suffix as – ene. e. g. α- Carotene, β- Carotene.

b) Xanthophyll: These are oxygenated (Oxygen derivative) hydrocarbon with empirical formula as C₄₀H₅₆O₂ and yellow in colour. The name of xanthophylls ends with suffix as – in. e.g. Lutein, Violoxanthin, Neoxanthin, Zeaxanthin, etc. Carotenes are present in PS-I and Xanthophylls in PS- II.

 

 

Functions of Carotenoids- 1) Protection of chlorophylls against photo-oxidation.

2) It absorbs and transfer light energy to the chlorophyll a for photosynthesis.

3) Phycobilins: Phycobilins are also tetrapyrol like chlorophylls but these four pyrol rings forms a straight chain. These are present in blue green algae and red algae. These are again two types as blue coloured C-phycocyanin and red coloured as r-phycoerythrin present in blue green algae and red algae, respectively.

 

 

 

 

In plants, chl. a and chl. b are main photosynthetic pigments while Carotenoids and Phycobilins are accessory photosynthetic pigments (the energy absorbed by them is not directly utilized in the photosynthesis).

 

 

 

Nature of light:

Light affects the photosynthesis in two ways as its quality (wavelength) and quantity (intensity). The visible light is only small portion of electromagnetic spectrum of light and it ranges from wave length 390 nm (violet) to 760 nm (red). Light travels in the form of a stream of discrete particles/ pocket of energy called photons. Thus the amount of energy in the photon is called as a quantum. The amount of energy present in the photon is inversely proportional to the wave length of light. Longer the wave length lesser the energy and vice versa.

Not all wave lengths of light are absorbed by the plant pigments. Maximum absorption of light occur in red and blue region and less absorption in yellow and orange region. Green light is mostly reflected. Thus the highest rate of photosynthesis occur in red and blue region of light.

 

 

Mechanism of Photosynthesis:

After performing many experiments F. F. Blackman (1905) observed that there are two types of reactions taking place during the photosynthesis.  Some reactions of photosynthesis takes place in presence of light or requires light hence called light reaction. These reactions are sensitive to light hence also known as photochemical reactions. However, some reactions of photosynthesis takes place in dark period or does not require light called Dark reaction or Blackman’s reaction. This reaction is sensitive to temperature hence known as thermochemical reaction.

A) Light reaction: (Hill reaction/Photochemical reaction)

This reaction takes place in presence of light and during which water is oxidized in the grana lamellae and forms the end products like NADPH₂, H₂O, ATP, and O₂. In the earlier days it was thought that the oxygen liberated during photosynthesis come from the CO₂. But latter on Hill (1937) and Ruben et. al. (1941) by using heavy isotope of oxygen (O18) proved that the oxygen released during photosynthesis is coming from H₂O molecules and not from CO₂. 

 

 

 

 

 

 

 

 

The light reaction involves two processes as a) Photolysis of water and b) Photophosphorylation.

a) Photolysis of water: The water molecules get oxidized or splitted into protons, electrons and release oxygen as byproduct.

 

 

 

The electrons are then transferred through electron transport system of photosynthesis and forms the ATP and reducing agents as NADPH₂. The energy and reducing agents are further used in dark reaction. For the transfer of electrons two photosystems are involved.

Photosystem:

The photosynthetic pigments present in grana lamellae are organized into an array (in definite system/structure) known as pigment system or photosystem. There are two photosystems which are named as photosystem-I (PS-I) and photosystem-II (PS-II). These photosystems are structurally and functionally distant from each other. Both of them contains chl. a, chl. b and carotenoids but their proportion varies.

Photosystem-I: The photosystem- I contains about 200 molecules of Chl. a, less amount of chl. b, carotenoids in the form of carotene, Iron, Copper, Plastocyanin and 1-2 molecules of ferredoxin. It also shows presence of cytochrome ‘b’, cytochrome ‘f’, ferredoxin reducing substance and P₇₀₀ as a reaction or trapping center. This chl. a molecule absorbs the longer wave length of light i.e. greater than 700 nm. PS- I is present only in the stroma lamellae and mainly functioning for up hilling of electrons to higher energy level by absorbing the radiant energy.

 

 

 

 

 

 

 

 

 

 

 

 

 

Photosystem-II: The major component of PS-II is chl. b, less amount chl. a, xanthophylls as violoxanthin and neo-xanthin. In addition to this, plastoquinone, Mn, Cl, cytochrome ‘b’ ₅₅₉ and cytochrome b₆. PS-II is present in grana lamellae and responsible for generating O₂ from water molecules and supplying electron to PS-I.

The electron from PS-II to PS-I are transferred through some intermediate like plastoquinone, Cytochrome ‘b₆’, cytochrome ‘f’ and plastocyanin and joins two light reactions.

B) Photophosphorylation:

The process of formation of ATP from ADP and inorganic phosphate by utilization of radiant energy is called photophosphorylation. OR The process of phosphorylation in presence of light is known as photophosphorylation.

Depending upon whether the electron lost by PS-I is cycled (come back to it) or not, there are two types of photophosphorylation. These are called 1) Cyclic and 2) Non-cyclic photophosphorylation.

1) Non-cyclic photophosphorylation:

In this type of photophosphorylation, both PS-I and PS-II are involved and the flow of electrons is unidirectional. During non-cyclic photophosphorylation, PS-II first absorbs the radiant energy and its pigments get excited. The excited chlorophyll molecules emit electrons from its outer most orbit to level of higher energy and undergoes oxidation. These electrons are first accepted by Phaeophytin and gets reduced. This reduced phaeophytin then oxidized by donating the electron to plastoquinone (PQ). Then the electron are transferred to cytochrome b6 and cytochrome f. During transfer of electrons from cytochrome b6 to cytochrome f some amount of energy is liberated that is utilized for synthesis of ATP from ADP and inorganic phosphate.

The cytochrome f then donate the electrons which are picked up by plastocyanin and finally by PS-I. The pigments of PS-I also absorb radiant energy and uphill or elevate them to higher level of energy. These electrons are then received by Ferredoxin reducing Substance (FRS) and passed on to Ferredoxin (Fd). The ferredoxin gets oxidized and electrons are transferred to coenzyme NADP and reduced to NADPH₂. This reaction is catalyzed by an enzyme Ferredoxin –NADP oxidoreductase. The electrons which are lost by PS-II always travels from one carrier to another carrier and cannot come back again to PS-II. Hence this electron transfer is called non-cyclic photophosphorylation.

 

 

 

The PS-II again accept new electron by the photolysis of water. During this, water is splitted into protons (H⁺). Electrons (e^-) and OH^- ions, which are further converted into H₂O molecules. The protons are also accepted by NADP and reduced to NADPH₂.

The end products of non-cyclic photophosphorylation are one molecule of ATP, NADPH₂, 2 molecules of H₂O and O₂.

2) Cyclic photophosphorylation:

The cyclic photophosphorylation involves only PS-I and takes place in presence of light, which have the wavelength greater than 680 nm. The chlorophyll molecules present in PS-I absorbs the sunlight energy, becomes exited and donate highly energized electrons. These electrons are first accepted by the Ferredoxin Reducing Substance (FRS) and then transferred to ferredoxin. The electrons from reduced ferredoxin are not passed on to NADP but transferred to cytochrome b₆. During this transfer, energy is liberated which is utilized for synthesis of first ATP. The electrons are then given to cytochrome f and during which again ATP is generated. The electron are ultimately come back to PS-I via Plastocyanin. The electron that is lost by PS-I come back to PS-I in a cyclic manner hence this is called cyclic photophosphorylation. The end product of cyclic photophosphorylation is only 2 molecules of ATP.

Thus totally 3 molecules of ATP and 1 molecule of NADPH₂ are the end products of light reaction. These are further utilized in the dark reaction. Therefore net gain of light reaction is only 2 molecules of H₂O and evolution of O₂.

 

 

B) Dark reaction: (Photosynthetic Carbon Reduction cycle/ Blackman’s reaction)

It take place in the stroma lamellae and independent (absence) of light hence known as Dark Reaction. During the dark reaction CO₂ is reduced to carbohydrates was initially identified by Blackman and therefore this process is also known as Blackman reactions. The photosynthetic CO₂ fixation in higher plants takes place by three different ways. These are classified on the basis of first stable photosynthetic product.  These three pathway are known as C₃, C₄ and CAM pathway and accordingly the plants are also classified as C₃, C₄ and CAM plants.

1) Calvin Cycle / C3 Pathway:

Calvin and his coworkers explained the reactions of this pathway by using unicellular alga Chlorella as photosynthetic system and radioactive isotope ¹⁴C as a tracer. The reactions of this pathway can be divided into 3 steps as a) Carboxylation b) Reduction and c) Regeneration of RuBP.

a) Carboxylaton: The reactions of this pathway are as follows-

1) The CO₂ is first accepted by RuBP (Ribulose 1, 5 biphosphate) and form a 6 carbon additive compound, which is unstable. This is then splitted into 2 molecules of 3-carbon containing compound i.e. 3- phosphoglyceric acid (3- PGA). This reaction is catalyzed by the enzyme Ribulose 1:5 biphosphate carboxylase / Rubisco).

2) The 3- PGA then undergoes phosphorylation and produces 1,3 diphosphoglyceric acid with the help of enzyme 3- phosphoglycerate kinase. In this reaction the end products of light reaction i.e. ATP is utilized.  

 

b) Reduction:

3) 1:3 diphosphoglyceric acid is then reduced by NADPH2 and forms 3-  Phosphoglyceraldehyde (3- PGAL). This reaction is catalyzed by 3-PGAL dehydrogenase enzyme.

4) The 3- PGAL is then undergoes process of isomerization into 3- dihydroxyacetone phosphate (3-DHAP) in presence of triose phosphate isomerase. These triose sugars are interconvertable.  

 5) Production of Fructose 1:6 diphosphate: One molecule of 3- PGAL combines with 3- DHAP to form fructose 1:6 diphosphate. The enzyme aldolase catalyses this reaction.

6) The fructose 1:6 diphosphate is then dephosphorylated by an enzyme fructose 1:6 diphosphatase and produces fructose 6 phosphate. The fructose 6 phosphate after isomerization produces glucose 6 phosphate by phosphofructose isomerase. The glucose 6 phosphate again undergoes dephosphorylation and forms glucose, which is further utilized for synthesis of starch as end products of photosynthesis.

c) Regeneration of RuBP:

7) One molecule of fructose 6 phosphate reacts with one molecule of 3- PGAL and produce one pentose sugar as Xylulose 5- phosphate and one tetrose sugar as erythrose 4- phosphate. This reaction is catalysed by an enzyme transketolase.  

8) Erythrose 4 phosphate combines with a molecule of 3-DHAP to form sedoheptulose 1:7 diphosphate. Enzyme transaldolase catalyses this reaction.

9)The sedoheptulose 1:7 diphosphate gets dephosphorylation and becomes sedoheptulose 7- phosphate with the help of enzyme diphosphatase.

10) Sedoheptulose 7- phosphate again combines with 3-DHAP with the help of enzyme transketolase and produces ribose 5- phosphate and xylulose 5- phosphate.

11) Both the pentose sugars are then readily isomerise to ribulose 5- phosphate in presence of enzyme ribose phosphate isomerase and epimerase, respectively.

12) Finally the ribulose 5 –Phosphate gets phosphorylation by using ATP and produces ribulose 1:5 diphosphate. This reaction is catalysed by ribulose phosphate kinase. The ribulose 1: 5 diphosphate again accept another CO₂ molecule and start new Calvin cycle.

Thus for reduction of one CO₂ molecules 2 molecules of NADPH₂ and 3 molecules of ATP are required.

 

 

 

 

 

 

2) C₄ pathway / HSK pathway:

Till 1965, it was believed that all land plants follows the C₃ pathway of photosynthesis. But in 1965, Kortschak et. al. working with ¹⁴CO₂ on sugarcane leaves found that a C₄ dicarboxylic acid i.e. Oxaloacetic acid (Malic acid or Aspartic acid) as first photosynthetic product instead of 3-PGA. Kortschak (1965) well established the C₄ pathway of photosynthesis in sugarcane, maize and number of grasses. Later on in many grasses and dicotyledon plants this pathway was investigated by Hatch and Slack (1967). They studied the kinetics of photosynthetic enzymes, products and cytology of these plants and finally proposed the C₄ pathway.  In honour of these scientists this pathway is known as Hatch, Slack and Kortschak (HSK) pathway.

1) In this pathway, first CO₂ acceptor is phosphoenol pyruvic acid (PEP), a 3 carbon containing compound and it produces Oxaloacetic acid (OAA). It is a dicarboxylic unstable 4- carbon containing acid. This reaction takes place in mesophyll cell and is catalysed by phosphoenol pyruvate carboxylase (PEP-case).

 

 

 

 

 

2) The OAA is then converted into either malic acid or aspartic acid. In some plants, the OAA enter in the chloroplast and with the help of malate dehydrogenase (MDH) and reducing agent NADPH₂, produces malic acid. While in some plants OAA is directly converted into aspartic acid by aspartate amino transferase enzyme.

3) The malic acid (Or aspartic acid) then transfer to bundle sheath cells where it undergoes oxidative decarboxylation and produces pyruvic acid and CO₂. In this reaction NADP gets reduced to NADPH₂ and enzyme is known as NADP dependent malic enzyme.

4) The pyruvic acid is transported back to the mesophyll cells where the pyruvate phosphate dikinase phosphorylate this to phosphoenol pyruvic acid. This enzyme requires ATP which is splitted into AMP and two molecules of inorganic phosphate (PPi).

5) The CO₂ released during oxidative decarboxylation of malic acid is then re-fixed in Calvin cycle in the bundle sheath cell chloroplast and produces starch as an end product.

            The leaf anatomy of C₄ plants is particularly interesting. The leaves of C₄ plants are characterized by a sheath of parenchymatous cells that are radially arranged around the vascular bundle. These vascular bundles are present in loosely arranged spongy parenchymatous mesophyll cells. This closed vascular bundle sheath arrangement is known as ‘Kranz anatomy’. That means the C₄ plants show dimorphic chloroplast i.e. two types of chloroplasts. The chloroplast of mesophyll cells is different structurally and functionally from those of bundle sheath cells chloroplasts. The difference between them is as follows------

 

Sr. No.	Character	Mesophyll cell chloroplast	Bundle sheath cell chloroplast
1	Shape	Slightly elongate	Spherical
2	Size	Smaller	Larger
3	Number per cell	More	Less
4	Ultra structure	Granal type, Both PS-I and PS- II present	Agranal type – Only PS- I is present.
5	Arrangement	Irregular	Regular – either  Centripetal or Centrifugal
6	First CO2 acceptor	PEP	RuBP
7	First stable product	Malic acid / Aspartic acid	3-Phosphoglyceric acid
8	Starch	Absent	Present

 

Significance of C₄ pathway: About 1500 plant species belonging to 18 families of angiosperms show C4 pathway of photosynthesis. These plants generally found in tropical and sub-tropical regions where intensity of light is very low CO₂ concentration and less availability of water hence plants change their photosynthetic pathway from C₃ to C₄ pathway.

3) Crassulacean Acid Metabolism (CAM- pathway):

In succulent plants, the accumulation of acids takes place during night period and which are again utilized during the next successive day period. This type of fluctuation in acidity status in the chlorophyllous tissue was first reported in members of family Crassulaceae and hence known as Crassulacean Acid Metabolism (CAM Pathway).

In 1978, Osmond believed that M. Grew (1662) was the first man to realize the acid accumulation in the succulent plant, Aloe vera. Hayne (1865) noted a remarkable taste difference in Bryophyllum leaves, as sour in the morning, tasteless in afternoon and bitter toward the evening. Later on number of scientist contributed to this and in 1978 Crassulacean Acid Metabolism pathway was fully established by Osmond, M. L kluge and Ting.

CAM pathway is known to occur among the members of 23 families of angiosperms including Crassulaceae, Agavaceae, Cactaceae, Liliaceae, Chenopodiaceae, Asclepidaceae, Euphorbiaceae, etc.

Metabolic Pathway:

The succulent xerophytic plants show some adaptations as to check the rate of transpiration the stomata remains closed during the day period and open during night period. The CO₂ fixation in these plants takes place during night period.  CAM plants are characterized by two metabolic processes which are separated by time. These two processes are – 1) Acidification – Dark process and 2) Deacidification- Light process.

1) Acidification – Dark process: During the night period, CO2 is first accepted by phosphoenol pyruvic acid (PEP) in the presence of enzyme PEP-case and form Oxaloacetic acid (OAA).

 

 

 

The OAA is then reduced to malic acid by the malate dehydrogenase enzyme. In this reaction NADPH2 acts as hydrogen donor and get oxidized as NADP. The synthesis and storage of malic acid takes place continuously during night period, hence the acidity status becomes higher early in the morning. This entire process is known as acidification.

2) Deacidification- Light process: During next daytime, the malic acid again enter in the cytoplasm where it undergoes oxidative decarboxylation and produces pyruvic acid and CO₂.  This reaction is catalysed by NADP dependent Malic enzyme. In this reaction hydrogen atoms are accepted by NADP and get reduced to NADPH2 and release of CO₂. The CO₂ is further enter into Calvin cycle and produces starch. Hence starch content is more late in the evening. The pyruvic acid is then utilized for re-synthesis of PEP.

The accumulation of malic acid occurs during night period and its utilization in the day period show diurnal fluctuation. In the same manner, the starch content also show diurnal fluctuation but it is completely reverse manner to that of acidity status.

The CAM plants exhibits the following 4 important characteristics as –

1) The stomata remain open during night period and closed during day period.

2) The CO₂ fixation during the night period.

3) The acidity status (malic acid) of photosynthetic tissue show diurnal fluctuation as it accumulates during night and utilized during day period.

4) Starch content also shows diurnal fluctuation but which is reverse to that of malic acid.

The main significance of CAM pathway is an adaptation to extreme xerophytic environmental conditions.

Significance of Photosynthesis:

1) Food: Photosynthesis is a food processing process. It provide food directly or indirectly to all organism. Thus this process has much significance as it sustain the life on the earth.

2) The photosynthesis is energy trapping process through which light energy enters in the biosphere.

3) Source of Oxygen: It is equally important for its byproduct as oxygen which is required for respiration of all living organisms.

4) It coupled with respiration and maintains the CO₂ and O₂ concentration in the atmosphere.

5) Fossil fuel: Coal and petroleum are two fossil fuels used for domestic and industrial purposes.

6) Photosynthesis acts as a bridge that connects the inorganic and organic world.

 

Short Form:

OOA  =  Oxalo Acetic Acid                                                   PEP = Phosphoenol pyruvate

3-PGA  = 3-Phosphoglyceric acid                                          PS-I = Photosystem – I

3-PGAL = 3-Phosphoglyceraldehyde                                    PS-II = Photosystem – II

PQ = Plastoquinone                                                                PC = Plastocyanin

NADP = Nicotine Amide Dinucleotide Phosphate               Fd = Ferredoxin

RuBp = Ribulose 1:5 biphosphate                                         ATP = Adenosine Tri Phosphate

ADP = Adenosine Di Phosphate                                            AMP = Adenosine Mono Phosphate  

RuBp-case (Rubisco) = Ribulose 1:5 Biphosphate Carboxylase.

3- DHAP = 3-Dihydroxy Acetone Phosphate.                      Cyt. b₆ = Cytochrome b₆

FRS = Ferredoxin Reducing Substance                                 Cyt. f = Cytochrome f