Unit – II

Photosynthesis:
Historical background, photosynthetic
pigments, Absorption of light, transfer of
light energy, photochemical reactions,
photosynthetic electron transport (PSI,
PSII, Q cycle),
CO2 reduction, photorespiration, Factors
affecting CO2 reduction.
 From where do plants obtain food?
• In the 1600s, Jan Van Helmont made a hypothesis.
He gathered that, in order for a tree to grow and gain mass, it must
obtain food from somewhere else. He carefully weighed a tree and
the soil that the he wanted to plant the tree in. Time passed, and
the tree grew. After five years, he reweighed the tree, which had
grown quite large, and then…wait for it…he weighed the soil. To his
surprise, the soil weighed basically the same amount every day.
Jan concluded that the tree must have gained the nutrients from
the water that he had added over the years.
• Jan Ingen-Housz
About 100 years later, another Jan, demonstrated that plants, well
actually, only the green parts like the stems and leaves, could
manufacture oxygen by using the Sun’s light and not its heat.
Furthermore, Jan proposed the novel idea that plants used carbon
dioxide (CO2) while producing oxygen (O2).
• In 1881, Theodor Engelmann discovered that photosynthesis
occurs in the chloroplast.
• Until the 1930s, the O2 produced from photosynthesis was
believed to originate from CO2. L.B. Niel, a graduate student at
Stanford University at the time, hypothesized that the
O2 produced during photosynthesis came from H2O rather
than from CO2.
• In 1941, two scientists named Samuel Ruben and Martin
Kamen did just that and showed that the O2 released during
photosynthesis came from H2O. Used isotopes!!!
• Melvin Calvin (1954)
Traced the path of CO2 assimilation in photosynthesis and
awarded the Nobel Prize in 1961.
THE SUN: MAIN SOURCE OF ENERGY
FOR LIFE ON EARTH
 Light

• Light has properties of both particles and waves

• Light is like a stream of particles called as photons
• The energy carried by a photon is called a quantum
• The energy of a photon depends on the frequency (ν) of light
as determined by Planck’s law

Where ‘h’ is the Planck constant (6.6x10-34 J.s), ‘c’ the velocity of
the light and ‘λ’ is the wavelength of light
Electromagnetic Spectrum and Visible
Light

Visible spectrum
 OVERVIEW OF PHOTOSYNTHESIS
• Photosynthesis is the process by which autotrophic
organisms use light energy to make sugar and oxygen
gas from carbon dioxide and water

• Makes organic molecules (glucose) out of inorganic

materials (carbon dioxide and water).
• It begins all food chains/webs. Thus all life is supported
by this process.
• It also makes oxygen gas!!
 Contd…

• Photosynthesis is the only process of biological

importance that can harvest the light energy
• Almost all plants are photosynthetic autotrophs, as are
some bacteria and protists
– Autotrophs generate their own organic matter through
photosynthesis
– Heterotrophs are consumers of these organic matter

Mosses, ferns, and Euglena Cyanobacteria

flowering plants
Photosynthesis can be oxygenic or anoxygenic
• Photosynthesis is a biological oxidation-reduction process
• CO2 is the electron acceptor and H2A is any reduced compound
which can donate electron to make carbohydrate molecule
(CH2O)
General reaction of photosynthesis can be written as -
CO2 + 2H2A (CH2O) + 2A + H2O

Oxygenic photosynthesis -
CO2 + 2H2O (CH2O) + O2 + H2O

Anoxygenic photosynthesis -
CO2 + 2H2S (CH2O) + 2S + H2O
Example: Purple sulfur bacteria
 Photosynthesis involves two processes
1. Light-dependent reaction
2. Light- independent reaction or Calvin cycle
H 2O CO2
Light
• The light Chloroplast
reactions Other
convert Chlorophyll materials
absorbs
solar energy Calvin
Cycle:
Light-
to chemical Dependent
Light-
Independent
Reactions H+ ions
energy ATP
Reactions

– Produce NADPH
ATP &
NADPH Sugar (Glucose)
O2 C6H12O6

• The Calvin cycle makes sugar from carbon dioxide

– ATP generated by the light reactions provides the energy for sugar
synthesis
– The NADPH produced by the light reactions provides the electrons for
the reduction of carbon dioxide to glucose
Light-dependent reaction or photochemical
reaction occurs in thylakoid of chloroplast
Chloroplast
LEAF CROSS SECTION MESOPHYLL CELL
LEAF

Mesophyll

CHLOROPLAST Intermembrane space

Outer
membrane

Granum Inner
membrane
Grana Stroma Thylakoid
Stromal Thylakoid
thylakoids compartment
 Absorption of light
• Absorption of light occurs by pigments present in
chloroplast
- Chlorophyll a
- Chlorophyll b
- Carotenoids Light Reflected
light

• Chlorophyll a is
considered as the main
pigment and chlorophyll b
and carotenoids are
considered as accessory
pigments Absorbed
light
• Chlorophyll pigments do
Transmitted
not absorb green light light
Chloroplast
 Contd…

Chlorophyll
absorbs
blue-violet
& red light
best
Chlorophyll molecules embedded as
complexes in the thylakoid membrane

• Located in the thylakoid

membranes
• Chlorophyll have Mg+ in
the center
• Chlorophyll pigments
harvest energy (photons)
by absorbing certain
wavelengths (blue-420 nm
and red-660 nm are most
important)
How does light energy convert in to chemical energy?
Light
Chlorophyll pigment Chlorophyll pigment* (Excited state)

Energy transfer- Resonance energy transfer

Photosynthesis takes
place in complexes
containing Light
harvesting Antennas and
photochemical reaction
centres
In 1932, Emerson and
Arnold Performed a key
experiment showing
cooperation of many
chlorophyll molecules in
energy conversion
during photosynthesis

Quantum yield (ɸ) = Number of photochemical products/Total

No of quanta absorbed
Oxygen evolving organisms have two photosystems
• By the late 1950s, Emerson measured the quantum
yield of photosynthesis as a function of wavelength
and revealed an effect known as Red drop.
• The quantum yield for the most of the ranges of
wavelengths at which chlorophyll absorbs light is
fairly constant. However the yield drops at far red
region of chlorophyll absorption (>680nm).
• Enhancement effect: Emerson used red and far red
light together and separately. The rate of
photosynthesis was grater when both the red and
far red light used together than the sum of the
individual lights.
Conclusion of Emerson Expt. - PSI and PSII
• During the light-dependent reaction,
there are two possible routes for
electron flow:

A. Cyclic Electron Flow

B. Noncyclic Electron Flow
 Cyclic Electron Flow
• Occurs in the thylakoid membrane
• Uses Photosystem I only
• P700 reaction center- chlorophyll a
• Uses Electron Transport Chain (ETC)
• Generates ATP only

ADP + P ATP
 Cyclic Electron Flow

Ferridoxin (Fd), an
iron containing
protein which acts as
an electron carrier

Plastoquinone (Pq)

Plastocyanin (Pc)

Pigments absorb light energy & excite e- of Chlorophyll a

to produce ATP
 Noncyclic Electron Flow
• Occurs in the thylakoid membrane
• Uses Photosystem II and Photosystem I
• P680 reaction center (PSII) - chlorophyll a
• P700 reaction center (PS I) - chlorophyll a
• Uses Electron Transport Chain (ETC)
• Generates O2, ATP and NADPH
Noncyclic Electron Flow
 Stroma

Lumen

2H2O + 2NADP+
Light
8 photons

2NADPH + 2H+ + O2
 Q cycle
• Plastoquinol (PQH2) formed in
PSII is oxidized by the
cytochrome b6f complex in a
series of steps like those of the
Q cycle in the cytochrome bc1
complex of mitochondria.
• One electron from PQH2 passes
to the Fe-S center of the Rieske
protein (purple), the other to
heme bL of cytochrome b6
(green).
• The net effect is passage of
electrons from PQH2 to the
soluble protein plastocyanin,
which carries them to PSI
 Chloroplast
Light
Stroma

Stack of NADP
thylakoids ADP
+P CO2 reduction
Light Calvin
reactions cycle

Sugar used for

 Cellular respiration
 Cellulose
 Starch
 Other organic compounds

• Light-independent reaction (Dark Reaction)

– Does not require light
– Calvin Cycle
• Occurs in stroma of chloroplast • C3 plants (80% of plants on earth)
• Requires CO2 • To produce glucose: it takes 6 turns
• Uses ATP and NADPH as fuel to run and uses 18 ATP and 12 NADPH.
• Makes glucose sugar from CO2 and Hydrogen
 Regulation of RubisCO
1st Enzyme in Calvin Cycle
Substrate/Product availability

Allosterically regulated by NADPH and ATP

Very Narrow pH optimum pH 8 / pH7

pH 8 or above, inactive at 7

Enzyme must be in
reduced form
30
 RubisCO
• The only enzyme that enables the fixation of
atmospheric CO2 for the formation of biomass
• A prerequisite for the existence of the present life
on Earth
• In plants and cyanobacteria, it consists of eight
identical large subunits (51–58 kDa) and eight identical
small subunits (12–18 kDa)
• With its 16 subunits, RubisCO is one of the largest
enzymes in nature
• In plants the genetic information for the large
subunit is encoded in the plastid genome and for
the small subunit in the nucleus
• Each large subunit contains one catalytic centre
 Photorespiration
• Photorespiratory pathway discovered in
1972 by the American scientist Edward
Tolbert
• Occurs on hot, dry, bright days (Stomata close)
• Fixation of O2 instead of CO2
• Produces 2-C molecules instead of 3-C sugar
molecules
• Produces no sugar molecules or no ATP
• In the course of photorespiration ATP is
consumed
RubisCO is capable of performing both
carboxylase & Oxygenase reaction
 glycolate phosphate
phosphatase

glycolate
oxidase

glutamate-glyoxylate
aminotransferase
 glycine decarboxylase
complex

Why are two other organelles besides the

chloroplasts involved in the recycling process of 2-
phosphoglycolate?
 UNIT III
Respiration in plants
Do plants breathe??
• Yes, plants require
oxygen for respiration
and they also give out
carbon dioxide.
• Plants unlike animals do
not have any specialized
organs for gaseous
exchange but they have
stomata and lenticels for
this purpose.
 Cellular Respiration
• Glucose occupies a central
position in the metabolism of
plants, animals, and many
microorganisms.

• It is relatively rich in potential

energy, and thus a good fuel.

• The complete oxidation of

glucose to carbon dioxide and
water proceeds with a standard
free-energy change of -2,840 Fate of glucose
kJ/mol.
 What happens during respiration
Glucose + oxygen Carbon dioxide + water + Energy
C6H12O6 + 6O2 6CO2 + 6H2O + Energy

The entire process of respiration can be divide in to 4 parts;

Glycolysis
Electrons carried in NADH

Electrons
Link reaction Pyruvic
acid
carried in
Glucose NADH and
Electron
Krebs FADH2
Glycolysis Transport
Cycle
Krebs cycle or TCA cycle Chain

Cytoplasm
Mitochondrion
Oxidative phosphorylation
(Electron Transport Chain)
 Glycolysis
• Glycolysis splits one molecule of Glucose (6-carbon
molecule) in to two molecules of Pyruvate (3-
carbon molecule)
• Glycolysis takes place in the cytoplasm of cells
• Glycolysis doesn’t need oxygen to take place and is
the first step of both aerobic and anaerobic
respiration
• Glycolysis was the first metabolic pathway to be
elucidated and is probably the best understood
• Glycolysis is an almost universal central pathway of
glucose catabolism – plants, animals,
microorganisms
 Glycolysis
a) Preparatory Phase and b) Pay off phase

Lehningers Principles of Biochemistry

Pay off phase
 Outputs of glycolysis

 2 ATPs (4 ATPs generated and 2 ATPs utilized)

 2 NADH
 2 H2O and 2H+
 2 Pyruvate (End product of glycolysis)
Possible fates of pyruvate formed during glycolysis

Some plant
tissues
 Regulation of Glycolysis
The most important control step of glycolysis is
the irreversible reaction catalysed by
Phosphofructo kinase (PFK)
• ATP/AMP: PFK allosterically inhibited by ATP
and the inhibition is reversed by AMP
• Citrate: PFK inhibited by high conc. of
citrate

Hexokinase: Inhibited by Glucose 6-P

 Link reaction
 Pyruvate is oxidized to Acetyl-CoA and CO2 by oxidative
decarboxylation
 Catalysed by pyruvate dehydrogenase complex consisting
of three enzymes
• Pyruvate dehydrogenase (E1)
• Dihydrolipoyl transacetylase (E2)
• Dihydrolipoyl dehydrogenase (E3)
 Requires five coenzymes
Citric acid cycle or tricarboxylic acid (TCA) cycle

 TCA cycle is also known as Krebs cycle after its

discoverer, Hans Krebs
 Eugene Kennedy and Albert Lehninger showed
in 1948 that, in eukaryotes, the entire set of
reactions of the citric acid cycle takes place in
mitochondria
 In eukaryotes it occurs in the matrix of
mitochondria and in bacteria it occurs in the
cytoplasm
 TCA cycle has 8 steps
Citric acid cycle
One molecule of pyruvate Two molecules of pyruvate
3 NADH 6 NADH
1 FADH 2 FADH
1 ATP 2 ATP
2 CO2 4 CO2
 Glycolysis

Link reaction

Krebs cycle or TCA cycle

Oxidative phosphorylation
(Electron Transport Chain)

Electron Transport Chain (ETC)

& Oxidative phosphorylation
 Mitochondrion

 Double membrane, with

inner membrane very
impermeable

 TCA occurs in the matrix

 ETC in the inner

membrane
 Electron Transport Chain (ETC)
• The vast majority of the ATP (90%) comes from the energy in
the electrons carried by NADH and FADH2.
• The energy in these electrons is used in the electron transport
chain to power ATP synthesis.

• Thousands of copies of the electron transport chain are found

in the extensive surface of the cristae (the inner membrane of
the mitochondrion).
• Electrons drop in free energy as they pass down the electron
transport chain.
Electron Transport Chain (ETC)

FADH2

Complex I – NADH dehydrogenase

II – Succinate dehydrogenase
III – Ubiquinone cytochrome c oxidoreductase
IV – Cytochrome oxidase
For each pair of electrons transferred to oxygen

 Electron transport chain sets up an H+ gradient

 The energy stored due to the difference in

proton concentration is called Proton Motive
Force (pmf).
 Unit – IV

Mechanism of ATP synthesis, substrate

level phosphorylation, chemiosmotic
mechanism (oxidative and
photophosphorylation), ATP synthase.
Synthesis and breakdown of triglycerides,
α-oxidation, β-oxidation, glyoxylate cycle
 Chemiosmotic theory and ATP synthesis
How is a concentration gradient of protons transformed in to ATP??

 The biochemical mechanism that couples proton flux with

phosphorylation is known as chemiosmotic model.

 The chemiosmotic model was proposed by Peter Mitchell.

 According to the model, the electrochemical energy inherent

in the difference in proton concentration and separation of
charge across the inner mitochondrial membrane—the
proton-motive force—drives the synthesis of ATP.

 Protons flow passively back into the matrix through a proton

pore associated with ATP synthase.
Chemiosmotic Model

ATP synthase
 ATP synthase
• This large enzyme complex of the inner mitochondrial
membrane catalyzes the formation of ATP from ADP and Pi
• Mitochondrial ATP synthase is an F-type ATPase similar in
structure and mechanism to the ATP synthases of chloroplasts
and eubacteria
• ATP synthase, also called Complex V.
ATP synthase has two distinct
components: F1, a peripheral membrane
protein, and Fo (o denoting oligomycin-
sensitive), which is integral to the
membrane
• F1, the first factor recognized as
essential for oxidative phosphorylation,
was identified and purified by Efraim
Racker and his colleagues in the early
1960s
 Lipids
• Lipids are chemically varied group of fatty substances
and are highly concentrated energy stores.
• Lipids are esters of long chain fatty acids and alcohol.
• They are water insoluble biomolecules but soluble in
organic solvents such as ether, benzene, chloroform
etc.
 Functions of lipid
• Lipids serve as fuel molecules and are highly
concentrated energy source.
• Lipids are constituents of cell membrane and regulate
membrane permeability.
• They protect internal organs by serving as insulating
materials and give shape and smoothness to the body.
• Serve as a source and carrier for fat soluble vitamins
• Essential fatty acids are useful for transport of
cholesterol, formation of lipoproteins etc.
• Cholesterol is a constituent of membrane structure
and it synthesizes bile acids, hormones, and vitamin D.
 Classification of lipids
Classification of lipids based on chemical composition
1. Simple lipids: Esters of fatty acids with alcohols
2. Complex lipids: Esters of fatty acids with alcohols
with additional groups phosphate, nitrogenous
base etc.
3. Derived lipids: Lipids that are obtained on
hydrolysis of simple and complex lipids. These lipids
contain glycerol and other alcohols
4. Miscellaneous lipids: These include compounds
which contain characteristics of lipids. They include
squalene, terpenes, hydrocarbons, carotenoids etc.
BILE SALTS
1. Simple Lipids
a. Fats & Oils
Three fatty acid molecules each in ester
linkage with a single glycerol
(Triacyglycerols)

b. Waxes
Long chain fatty acid (C14-C36) esters of
long chain alcohols (C16-C30)
Structure of Triacylglycerol
 Saturated fatty acid
Most Commonly occurring are
• Myristic Acid (14:0)
• Palmitic Acid (16:0)
• Stearic Acid (18:0)

Since the carboxylic group is ionized at

physiological pH, they exist as carboxylate ion
e.g. palmitate
CH3 (CH2)14-COOH CH3(CH2)14-COO- + H+
pKa 4.8
 Unsaturated fatty acids
Contain carbon to carbon double bonds.
MUFA
• Mono-unsaturated (1 double bond)
OLEIC ACID 18:1 ( high concentrations in olive oil )
PUFA
• Polyunsaturated (> 1 double bond )
LINOLEIC ACID, 18:2
LINOLENIC ACID, 18:3
ARACHIDONIC ACID 20:4
 Positional and Geometric isomerism in unsaturated
fatty acids
• cis configuration mostly present in naturally occurring fatty
acids
• cis double bond causes a bend. Therefore the hydrocarbon
chain cannot be packed as tightly as in trans fatty acids
• Melting points of fatty acids with cis double bonds are lower
than the corresponding trans fatty acids
 Biosynthesis of fatty acids in plants
• Lipids are membrane constituents and function as
carbon stores.
• Lipids are subdivided into glycerolipids,
sphingolipids, and steroids.
• Glycerolipids are fatty acid esters of glycerol.
• These lipids have fatty acid chains as hydrophobic
tail.
• Triacylglycerols are primarily present in seeds but
also in some fruits such as olives or avocados.
• Triacylglycerols are deposited in oil bodies, also
termed oleosomes or lipid bodies.
• A variety of oil body proteins (oleosines,
caloleosines, steroleosines) are anchored to the
lipid monolayer and catalyze the mobilization of
fatty acids from the triacylglycerol store during
seed germination.
• Triacylglycerols have an advantage over
carbohydrates as storage compounds, because
their weight to carbon content ratio is much
lower.
• Carbon stored as fat in the seed requires less
than half the weight as when it is stored as
starch.
• Low seed weight is advantageous for dispersal.
 The de novo synthesis of fatty acids
takes place in the plastids
• Synthesis of fatty acids occurs only for the
cell’s own requirements, except in seeds and
fruits.
• Plants are not capable of long-distance fatty
acid transport.
• Synthesis of fatty acids always occurs in the
plastids: in the chloroplasts of green cells and the
leucoplasts and chromoplasts of non-green cells.
• Modification of fatty acid chain may occur in
endoplasmic reticulum. E.g. Introduction of double
bonds to fatty acid chain by desaturase enzyme.
 Acetyl-CoA is a precursor for the
synthesis of fatty acids
• Acetyl CoA can be provided in two different ways.
• Plastids contain a pyruvate dehydrogenase complex, by
which pyruvate is oxidized to acetyl CoA.
• Chloroplasts contain a high activity of acetyl CoA synthetase,
which can convert acetate upon consumption of ATP to
acetyl CoA.
• In chloroplasts,
photosynthesis
provides the
NADPH required
for the synthesis
of fatty acids
• Fatty acid synthesis starts with the carboxylation of
acetyl CoA to malonyl CoA by acetyl CoA carboxylase.
• CoA is exchanged by acyl carrier protein (ACP).
• The enzyme β-
ketoacyl-ACP
synthase III (KAS
III) catalyzes the
condensation of
acetyl CoA with
malonyl-ACP.

CCRDR reactions
of fatty acid
synthesis
• β-Ketoacyl-ACP is reduced by NADPH to β-D-hydroxyacyl-ACP
• The carbon-carbon double bond formed is reduced by NADPH to
produce acyl ACP

• The product is a
fatty acid that has
been elongated by
two carbon atoms
until 16C palmitoyl

Net reaction of fatty acid synthesis:

8 Acetyl-CoA + 14 NADPH + 7 ATP
Palmitate + 14 NADP + 7 CoA + 7 ADP + 7Pi
 Acetyl CoA carboxylase is the first
enzyme of fatty acid synthesis
• Catalyzed first committed step in fatty acid synthesis
• Multienzyme complex consisting of three subunits:
1. Biotin carboxyl carrier protein
2. Biotin carboxylase
3. Carboxyl transferase
• The acetyl CoA carboxylase multienzyme complex in
the stroma of plastids resembles the acetyl CoA
carboxylase in cyanobacteria and other bacteria, and
is referred to as the prokaryotic form
 Contd….

• Eukaryotic form is also present in

chloroplast and cytoplasm of plants
with exceptions.
 Contd….

• In Gramineae (grass family), including the various

species of cereals, the prokaryotic form is not
present. In these plants, the multifunctional
eukaryotic acetyl CoA carboxylase is located in the
cytosol as well as in the chloroplasts.
• The eukaryotic acetyl CoA carboxylase is
inhibited by Diclofop methyl.
• In Gramineae, Diclofop methyl severely impairs
lipid biosynthesis in plastids.
• Diclofop methyl (trade name Hoe-Grass, Bayer
Crop Science) is therefore used as selective
herbicides.
 Regulation of Acetyl CoA carboxylase
• Acetyl CoA carboxylase, the first enzyme of fatty
acid synthesis, is an important regulatory enzyme.
• NADPH required by fatty acid synthesis is provided by
photosynthesis.
• In chloroplasts, the enzyme is fully active only during
illumination and is inhibited during darkness.
• The acetyl CoA carboxylase is reductively activated by
thioredoxin.
• Activity is further enhanced by the increase of
the pH and the Mg2+ concentration in the stroma.
 Beta oxidation of fatty acid
• Oxidation of fatty acids in to acetyl-CoA is called
β-oxidation
• β-oxidation proceeds in specialized peroxisomes
called glyoxysomes

Stored Triacylglycerols in seeds

Lipases

Glycerol 3-phosphate Glycerol + Fatty acids

β-oxidation
Glycolytic pathway Acetyl-CoA TCA cycle
Biosynthesis
Beta oxidation of fatty acid
Difference in biosynthesis and oxidation of Fatty acids

NADPH
 Glyoxylate cycle
• Conversion of Acetyl-CoA in to succinate for synthesis of hexoses
is called glyoxylate cycle
• Glyoxylate cycle enables plants to synthesize glucose from lipids
via gluconeogenesis
• Glyoxylate cycle occurs in glyoxysomes with two special enzymes
i) Isocitrate lyase
Converts isocitrate in to
succinate and glyoxylate

ii) Malate synthase

Converts glyoxylate in to
malate
 Contd…..

Malate synthase

Isocitrate lyase

Succinate

Mitochondria
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