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Module 4 (Metabolism in Cell)

Cellular Metabolism

4.1. Cellular metabolism

Self-Test
1. Where do cells obtain energy and why do they need it?
2. How cells gain energy?
3. What is metabolism and metabolic pathway?

Living cells are in a constant activity. Macromolecules are assembled and broken down, substances are
transported across cell membranes, and genetic instructions are transmitted. All of these cellular activities
require energy.

Living organisms are unique in that they can extract energy from their environments and use it to carry out
activities such as movement, growth and development, and reproduction. But the basic question is how
living organisms or, their cells extract energy from their environments, and how cells use this energy to
synthesize and assemble the components from which the cells are made. The answers to these questions
lie in the enzyme-mediated chemical reactions that take place in living matter (metabolism). Hundreds of
coordinated, multistep reactions, fueled by energy obtained from nutrients and/or solar energy, ultimately
convert readily available materials into the molecules required for growth and maintenance.

Metabolism is thus the sum of chemical reactions that takes place within each cell of a living organism and
that provides energy for vital processes and synthesizing of new organic materials. Broadly, these reactions
can be divided into catabolic reactions that convert nutrients to energy and anabolic reactions that lead
to the synthesis of larger biomolecules.

In metabolism, a series of chemical reactions in which the product of one reaction is the substrate for the
next reaction is called a metabolic pathway. Catabolic pathways release energy by breaking down larger
molecules into smaller molecules. Anabolic pathways use the energy released by catabolic pathways to
build larger molecules from smaller molecules, insuring the continual flow of energy within an organism.
The reactants and products of these chemical reactions are metabolites. The major classes of metabolites
include proteins, carbohydrates, nucleotides, lipids, coenzymes, and cofactors. These classes of
compounds encompass an enormous diversity of molecular structures, physicochemical properties,
functions, and abundances. Due to the analytical challenges posed by this diversity, most studies require
division into subsets of metabolites. The most common criteria for this distinction rely on the hydrophilic
(polar) and hydrophobic (nonpolar) nature of metabolites. Polar metabolites are soluble in aqueous
solutions and include most sugars, purines and pyrimidines, nucleotides and nucleosides, acyl carnitines,
organic acids, hydrophilic acids, amino acids, and phosphorylated compounds. These metabolites include
most of the reactants and products involved in cellular respiration (e.g., glycolysis, the TCA cycle, or in
other pathways) and in the production of building blocks for synthesis of large biopolymers such as DNA,
RNA, proteins, and oligosaccharides. The nonpolar or hydrophobic metabolites are commonly lipids. These
metabolites function in energy storage, membrane structure, and signal transduction.
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At the cellular level of organization, the main chemical processes of all living matter are similar, if not
identical. This is true for animals, plants, fungi, or bacteria; where variations occur (such as, the secretion of
antibodies by some molds), the variant processes are but variations on common themes. Thus, all living
matter is made up of large molecules called proteins, which provide support and coordinated movement, as
well as storage and transport of small molecules, and, as catalysts, enable chemical reactions to take place
rapidly and specifically under mild temperature, relatively low concentration, and neutral conditions (i.e.,
neither acidic nor basic).

The chemical reactions that take place in living cells are similar as well. Green plants use the energy of
sunlight to convert water (H2O) and carbon dioxide (CO2) to carbohydrates (sugars and starches), other
organic (carbon-containing) compounds, and molecular oxygen (O2). The process of photosynthesis
requires energy, in the form of sunlight, to split one water molecule into one-half of an oxygen molecule
(O2; the oxidizing agent) and two hydrogen atoms (H; the reducing agent), each of which dissociates to
one hydrogen ion (H+) and one electron.
Through a series of oxidation-reduction reactions, electrons (denoted e−) are transferred from a donating
molecule (oxidation), in this case water, to an accepting molecule (reduction) by a series of chemical
reactions; this “reducing power” may be coupled ultimately to the reduction of carbon dioxide to the level of
carbohydrate. In effect, carbon dioxide accepts and bonds with hydrogen, forming carbohydrates (Cn [H2O]
n). Living organisms that require oxygen reverse this process: they consume carbohydrates and other
organic materials, using oxygen synthesized by plants to form water, carbon dioxide, and energy. The
process that removes hydrogen atoms (containing electrons) from the carbohydrates and passes them to
the oxygen is an energy-yielding series of reactions. In plants, all but two of the steps in the process that
converts carbon dioxide to carbohydrates are the same as those steps that synthesize sugars from simpler
starting materials in animals, fungi, and bacteria. Similarly, the series of reactions that take a given starting
material and synthesize certain molecules will be used in other synthetic pathways are similar, or identical,
among all cell types. For instance, from a metabolic point of view, the cellular processes that take place in a
lion are only marginally different from those that take place in a dandelion.

4.2. Enzymes and their role in metabolism

Most chemical reactions within cells do not occur spontaneously. Instead, they need a catalyst to get them
started.

Why do you think this is so?

In many cases, heat may be a catalyst, but this is inefficient because heat cannot be applied to molecules
in a controlled fashion. Thus, most chemical reactions require biological catalyst called enzymes.

What are enzymes and why they are needed?

Enzymes are protein catalysts that speed biochemical reactions by facilitating the molecular
rearrangements that support cell function. Enzymes speed up (catalyze) chemical reactions; in some
cases, enzymes can make a chemical reaction millions of times faster than it would have been without it.
Almost all metabolic processes in the cell need enzyme catalysis in order to occur at rates fast enough to
sustain life.
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Enzymes bind with particular reactants until the chemical reaction occurs, then free themselves. Thus, at
any given time, the numerous pathways involved in building up and breaking down cellular components
must be monitored and balanced in a coordinated fashion.
To achieve this goal, cells organize reactions into various enzyme-powered pathways.

How do enzymes speed up chemical reactions?

Enzymes speed up reactions by lowering activation energy. Many enzymes change shape when substrates
bind. This is termed "induced fit", meaning that the precise orientation of the enzyme required for catalytic
activity can be induced by the binding of the substrate.

What is activation energy?

4.2.1 Chemical nature and classification of enzymes
All known enzymes are proteins with the exception of recently discovered RNA enzymes. Some enzymes
may additionally contain a non-protein group. Enzymes are high molecular weight compounds made up
principally of chains of amino acids linked together by peptide bonds.

Many enzymes require the presence of other compounds (cofactors) before their catalytic activity can be
exerted. This entire active complex is referred to as the holoenzyme; i.e., Apo enzyme (protein portion)
plus the cofactor (coenzyme, prosthetic group or metal-ion activator). Thus, on the basis of differences in
chemical nature, the enzymes may be described as follows:

 Simple enzymes: Simple enzymes are made up of only protein (polypeptide). They contain no
chemical groups other than amino acid residues. Digestive enzymes such as pepsin, and trypsin
are of this nature.

 Conjugate Enzymes: It is an enzyme which is formed of two parts – a protein part called Apo
enzyme (e.g., flavoprotein) and a non-protein part named cofactor. The complete conjugate
enzyme, consisting of an Apo enzyme and a cofactor, is called holoenzyme. There can be an
enzymatic activity only when both components
(Apo enzyme and cofactor) are present together. The cofactor is sometimes a simple divalent metallic ion
(e.g. Ca, Mg, Zn, Co, etc.), and sometimes a non-protein organic compound. However, some enzymes
require both kinds of cofactors. If the cofactor is firmly bound to the Apo enzyme, it is called prosthetic
group. For example, cytochromes are the enzymes that possess porphyrins as their prosthetic groups. If,
instead of being more or less permanently bound to the Apo enzyme the cofactor attaches itself to the Apo
enzyme only at the time of reaction, it is called a coenzyme.

 Metalloid-enzymes: The metal cofactors involved in enzyme reactions are monovalent (K+) and
divalent cations (Mg++, MN++ and Cu++). These may be loosely held by the enzyme, or as in some
cases, go into the composition of the molecule itself. If the metal forms part of the molecule, as iron
of hemoglobin or cytochrome is, the enzymes are called metallo-enzymes.

 Isozymes (Isozymes): At one time it was believed that an organism has only a single enzyme for
a given step of a metabolic reaction. It was later discovered that a substrate may be acted upon by
a number of variants of an enzyme producing the same product.
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Classes of enzymes based on the substrate they act up on

Enzymes can be classified based on different criteria of which classification based on the substrate they
acted up on is the most common (Table 4.1).

Table 4.1. Major classes of Enzymes

4.2.2 Mechanisms of enzyme action

Mechanisms of enzyme catalysis vary, but are all similar in principle to other types of chemical catalysis in
that the crucial factor is a reduction of energy barrier(s) separating the reactants from the products. The
reduction of activation energy increases the fraction of reactant molecules that can overcome this barrier
and form the product. An important principle is that since they only reduce energy barriers between
products and reactants, enzymes always catalyze reactions in both directions, and cannot drive a reaction
forward or affect the equilibrium position.

An enzyme attracts substrates to its active site, catalyzes the chemical reaction by which products are
formed, and then allows the products to dissociate (separate from the enzyme surface). The combination
formed by an enzyme and its substrates is called the enzyme– substrate complex. The substrates are
attracted to the active site by electrostatic and hydrophobic forces, which are called noncovalent bonds
because they are physical attractions and not chemical bonds.

Do you recall enzyme models? Discuss on it in detail.

4.2.3 Factors affecting enzymatic activities
The activity of an enzyme is affected by its environmental conditions such as temperature and pH.
Changing these alter the rate of reaction caused by the enzyme. In nature, organisms adjust the conditions
of their enzymes to produce an optimum rate of reaction, where necessary, or they may have enzymes
which are adapted to function well in extreme conditions where they live.

Temperature
Increasing temperature increases the kinetic energy that molecules possess. In a fluid, this means that
there are more random collisions between molecules per unit time. Since enzymes catalyze reactions by
randomly colliding with substrate molecules, increasing temperature increases the rate of reaction, forming
more product.
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However, increasing temperature also increases the vibrational energy that molecules have, specifically
(in this case enzyme molecules), which puts strain (damage) on the bonds that hold them together. As
temperature increases, more bonds, especially the weaker hydrogen and ionic bonds, will break as a result
of this strain. breaking bonds within the enzyme will cause the active site to change shape. This change in
shape means that the Active Site is less complementary to the shape of the Substrate, so that it is less
likely to catalyze the reaction. Eventually, the enzyme will become denatured and will no longer function.
The temperature at which the maximum rate of reaction occurs is called the enzyme‟s Optimum
Temperature.
This is different for different enzymes.

Activity
1. Graphically, show the relationship between enzyme activity and temperature.
2. What is the optimum temperature for most enzymes in the human body?

pH - Acidity and Basicity pH is a measure of the hydrogen ion (H+) concentration, and therefore a good
indicator of the hydroxide ion (OH-) concentration. Lower pH values mean higher H+ concentrations and
lower OH- concentrations. H+ and OH- ions are charged and therefore interfere with hydrogen and ionic
bonds that hold together an enzyme, since they will be attracted or repelled by the charges created by the
bonds. This interference causes a change in shape of the enzyme, and importantly, its active site.
Different enzymes have different optimum pH values. This is the pH value at which the bonds within them
are influenced by H+ and OH- ions in such a way that the shape of their active site is the most
complementary to the shape of their substrate. At the optimum pH, the rate of reaction is at an optimum.
Any change in pH above or below the optimum will quickly cause a decrease in the rate of reaction, since
more of the enzyme molecules will have active sites whose shapes are not (or at least are less)
complementary to the shape of their substrate.

Small changes in pH above or below the optimum do not cause a permanent change to the enzyme, since
the bonds can be reformed. However, extreme changes in pH can cause enzymes to denature and
permanently lose their function. Enzymes in different locations of our body have different optimum pH
values since their environmental conditions may be different. For example, the enzyme pepsin functions
best at around pH-2 and is found in the stomach, which contains hydrochloric acid.

Because enzymes are sensitive to changes in acidity, most living systems are highly buffered; i.e., they
have mechanisms that enable them to maintain a constant acidity.

Activity
What do you know about a buffer and its importance in the body of organisms?

Substrate and enzyme concentration

Changing the enzyme and substrate concentrations affect the rate of reaction of an enzymecatalysed
reaction. Controlling these factors in a cell is one way that an organism regulates its enzyme activity and so
its metabolism. Changing the concentration of a substance only affects the rate of reaction if it is the limiting
factor. If it is the limiting factor, increasing concentration will increase the rate of reaction up to a point, after
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which any increase will not affect the rate of reaction. This is because it will no longer be the limiting factor
and another factor will be limiting the maximum rate of reaction. As a reaction proceeds, the rate of reaction
will decrease, since the substrate will get used up.

Substrate concentration
Increasing substrate concentration increases the rate of reaction. This is because more substrate
molecules will be colliding with enzyme molecules, so more product will be formed.
However, after a certain concentration, any increase will have no effect on the rate of reaction, since
substrate concentration will no longer be the limiting factor. The enzymes will effectively become saturated,
and will be working at their maximum possible rate.

Enzyme concentration
Increasing enzyme concentration will increase the rate of reaction, as more enzymes will be colliding with
substrate molecules. However, this too will only have an effect up to a certain concentration, where the
enzyme concentration is no longer the limiting factor.

Activity
Show graphically the relationships between enzyme activity and substrate concentration as well as enzyme
concentration

Enzyme inhibitors
Enzyme activity can be inhibited in various ways. Inhibition could be reversible or irreversible.

Reversible inhibition
Competitive inhibition: occurs when molecules very similar to the substrate molecules bind to the active
site and prevent binding of the actual substrate. Penicillin, for example, is a competitive inhibitor that blocks
the active site of an enzyme that many bacteria use to construct their cell walls.

Noncompetitive inhibition: occurs when an inhibitor binds to the enzyme at a location other than the
active site. In some cases of noncompetitive inhibition, the inhibitor is thought to bind to the enzyme in such
a way as to physically block the normal active site. In other instances, the binding of the inhibitor is believed
to change the shape of the enzyme molecule, thereby deforming its active site and preventing it from
reacting with its substrate. This latter type of noncompetitive inhibition is called allosteric inhibition; the
place where the inhibitor binds to the enzyme is called the allosteric site. Frequently, an end-product of a
metabolic pathway serves as an allosteric inhibitor on an earlier enzyme of the pathway. This inhibition of
an enzyme by a product of its pathway is a form of negative feedback.

Activators: Allosteric control can involve stimulation of enzyme action as well as inhibition. An activator
molecule can be bound to an allosteric site and induce a reaction at the active site by changing its shape to
fit a substrate that could not induce the change by itself. Common activators include hormones and the
products of earlier enzymatic reactions. Allosteric stimulation and inhibition allow production of energy and
materials by the cell when they are needed and inhibit production when the supply is adequate.

Irreversible inhibition
Irreversible inhibitors usually covalently modify an enzyme, and inhibition can therefore not be reversed.
Irreversible inhibitors often contain reactive functional groups. Irreversible inhibition is different from
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reversible enzyme inactivation. Irreversible inhibitors are generally specific for one class of enzyme and do
not inactivate all proteins; they do not function by destroying protein structure but by specifically altering the
active site of their target

Activity
1.Describe other types of inhibition.
2.Relate the concept of irreversible enzyme inhibition to drug discovery and food poisoning.
3.Explain how painkillers like ibuprofen relieved you from pain.
4.What is the difference between endothermic and exothermic reaction?
5.What is meant by a catalyst?
6. Why are several enzymes needed in a typical metabolic path way?
7. Which of the three (coenzyme, cofactor, Apo enzyme) is a protein?
8.Descibe three factors that that influence enzymatic activity.

4.3 Bioenergetics and biosynthesis

4.3.1. Cellular respiration
Most living organisms obtain energy by breaking down organic molecules (catabolism) during cellular
respiration. The function of cellular respiration is to harvest electrons from carbon compounds, such as
glucose, and use that energy to make Adenosine Tri Phosphate (ATP). ATP is used to provide immediate
energy for cells to do work. This catabolic process can be divided into 3 phases.

Phase I - Breakdown of large complex biomolecules like polysaccharides, proteins and lipids into
their respective building blocks (hydrolysis). The chemical reactions occurring during this stage do
not release much energy.
Phase II - These building blocks are usually oxidized to a common intermediate, acetyl - CoA.
Additionally, pyruvate or other citric acid cycle intermediates may also be formed (in glycolysis and
other pathways).
Phase III – This consists of the citric acid cycle (i.e. oxidation of acetyl - CoA to CO2, formation of
NADH and FADH2) followed by electron transport and oxidative phosphorylation. Energy released
by electron transport to O2 is coupled to ATP synthesis. This cycle is responsible for the release of
much energy (TCA cycle and ETC).

Cellular respiration occurs in two main parts: glycolysis and aerobic respiration. The first stage, glycolysis,
is an anaerobic process. Anaerobic metabolic processes do not require oxygen. Aerobic respiration
includes the Krebs cycle and electron transport chain and is an aerobic process. Aerobic metabolic
processes require oxygen.

4.3.1.1. Glycolysis: anaerobic respiration

Glucose is a key metabolite in metabolism. Various pathways that are concerned with the utilization,
storage, and regeneration of glucose exist. Glycogen is a polymeric storage form of glucose in human and
it is most abundant in the liver and in striated muscle, although some is found in other tissues also.
Glycogen is synthesized when glucose supply is high, and its degradation helps to maintain the blood
glucose level when we are fasting. When glycogen is depleted, more glucose is synthesized from scratch in
gluconeogenesis. This pathway‟s most important substrates are amino acids, which are obtained either
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from a protein-rich diet-for example, during fasting on meat exclusively-or, during starvation, from
breakdown of cellular protein, mainly in skeletal muscle. Gluconeogenesis occurs in the liver and in the
kidneys.

Self-test
1. Define terms (glycolysis and gluconeogenesis)
2. Why glucose is more preferred by cells for respiration than other carbohydrates/lipids?
3. How is anaerobic respiration differing from aerobic respiration?
4.What are the three basic metabolic stages in respiration?

The first step in the degradation of glucose is glycolysis, which breaks down glucose to pyruvate. The main
purpose of glycolysis is the generation of energy (ATP). A modest amount of ATP is produced in glycolysis
directly, but much more ATP is formed downstream of glycolysis through the complete oxidation of
pyruvate. Glycolysis is the most common pathway for glucose degradation to pyruvate and is found in
animals, plants and microorganism. This pathway is used by anaerobic as well as aerobic organisms. The
process takes place in the cytoplasm of prokaryotes and eukaryotes and does not require oxygen. Under
aerobic conditions, most of the pyruvate formed in glycolysis undergoes complete oxidative degradation to
CO2 and H2O.

Pyruvate intended for complete degradation is transported to the mitochondria, where it is decarboxylase to
Acetyl-CoA by pyruvate dehydrogenase. Acetyl-CoA is completely degraded in the citric acid cycle (or
tricarboxylic acid cycle; TCA cycle for short). The “H2” that is produced here is not gaseous but bound to co-
substrates, as NADH and FADH2, which is subsequently oxidized in the respiratory chain.

If glucose is available in excess of immediate needs and glycogen is already stocked up to capacity, it
will still be broken down by glycolysis and pyruvate dehydrogenase to acetyl-coA. However, Acetyl-
CoA will then not be oxidized, but it will instead be used for fatty acid synthesis; the fatty acids are
converted to triacylglycerol. Fatty acid synthesis occurs in the cytosol of cells in the liver and fat tissue.

Activity
1. Which phases are regarded as the preparatory (or investment) phase? Why?
2. What is the purpose of each step?
3. What is the fate of pyruvate?

Glycolysis involves ten enzymatic reactions as described below.

The first five are preparatory phases or investment phase. Here, glucose is phosphorylated, rearranged and
phosphorylated again, with the two phosphate groups coming from ATP.

1. The phosphorylation of glucose at carbon-6 by hexokinase forming glucose 6phosphate (G6P).

 This reaction consumes ATP, but it acts to keep the glucose concentration low, promoting
continuous transport of glucose into the cell through the plasma membrane transporters. In
addition, it blocks the glucose from leaking out – the cell lacks transporters for G6P, and free
diffusion out of the cell is prevented due to the charged nature of G6P. Glucose may
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alternatively be formed from the phosphorolysis or hydrolysis of intracellular starch or

glycogen.
 In animals, an isozyme of hexokinase called glucosidase is also used in the liver, has a much
lower affinity for glucose, and differs in regulatory properties. The different substrate affinity
and alternate regulation of this enzyme are a reflection of the role of the liver in maintaining
blood sugar levels.
2. The conversion of glucose-6-phosphate(G6P) to fructose-6-phosphate(F6P) by phosphohexose
isomerase
 The change in structure is an isomerization, in which the G6P has been converted to F6P. This
reaction is freely reversible under normal cell conditions. However, it is often driven forward
because of a low concentration of F6P, which is constantly consumed during the next step of
glycolysis. Under conditions of high F6P concentration, this reaction readily runs in reverse.
This phenomenon can be explained through Le Chatelier's Principle, ''Isomerization to a keto
sugar is necessary for carbanion (negative charge of carbon that is stable) stabilization in the
fourth reaction step (below)''.
3. The phosphorylation of fructose-6-phosphate to the 1,6-
bisphosphate by phosphofructokinase,
 The energy expenditure of another ATP in this step is justified in 2 ways: The glycolytic
process (up to this step) becomes irreversible, and the energy supplied destabilizes the
molecule. Because the reaction catalyzed by Phosphofructokinase 1 (PFK-1) is coupled to the
hydrolysis of ATP (an energetically favorable step) it is, in essence, irreversible, and a different
pathway must be used to do the reverse conversion during gluconeogenesis. This makes the
reaction a key regulatory point. This is also the rate-limiting step.
 The second phosphorylation event is necessary to allow the formation of two charged groups
(rather than only one) in the subsequent step of glycolysis, ensuring the prevention of free
diffusion of substrates out of the cell.
 The same reaction can also be catalyzed by pyrophosphate-dependent phosphofructokinase
(PFP or PPi-PFK), which is found in most plants, some bacteria, archaea, and protests, but not
in animals. This enzyme uses pyrophosphate (PPi) as a phosphate donor instead of ATP. It is
a reversible reaction, increasing the flexibility of glycolytic metabolism.
4. The cleavage of fructose-1,6-bisphosphate by aldolase. This yields two different products,
dihydroxyacetone phosphate and glyceraldehyde-3-phosphate,
 Destabilizing the molecule in the previous reaction allows the hexose ring to be split by
aldolase into two triose sugars: dihydroxyacetone phosphate (a ketose), and glyceraldehyde 3-
phosphate (an aldose). There are two classes of aldolases: class I aldolases, present in
animals and plants, and class II aldolases, present in fungi and bacteria; the two classes use
different mechanisms in cleaving the ketose ring.
 Electrons delocalized in the carbon-carbon bond cleavage associate with the alcohol group.
The resulting carbanion is stabilized by the structure of the carbanion itself via resonance
charge distribution and by the presence of a charged ion prosthetic group.
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5. The isomerization of dihydroxyacetone phosphate to a second molecule of glyceraldehyde-3-

phosphate by triose phosphate isomerase,
Triose phosphate isomerase rapidly interconvert dihydroxyacetone phosphate with
glyceraldehyde 3-phosphate (GADP) that proceeds further into glycolysis. This is
advantageous, as it directs dihydroxyacetone phosphate down the same pathway as
glyceraldehyde 3-phosphate, simplifying regulation.

Remember!!
The rest five are pay-off phases which are characterized by a net gain of the energy-rich molecules ATP
and NADH. Since glucose leads to two triose sugars in the preparatory phase, each reaction in the pay-off
phase occurs twice per glucose molecule. This yields 2 NADH molecules and 4 ATP molecules, leading to
a net gain of 2 NADH molecules and 2 ATP molecules from the glycolytic pathway per glucose. Along the
way, the two molecules are rearranged into pyruvate.

6. The dehydrogenation and concomitant phosphorylation of glyceraldehyde-3-phosphate to 1,3-bis-

phosphoglycerate by glyceraldehyde-3-phosphate dehydrogenase,
 The aldehyde groups of the triose sugars are oxidized, and inorganic phosphate is added to
them, forming 1,3-bisphosphoglycerate.
 The hydrogen is used to reduce two molecules of NADH, a hydrogen carrier, to give NADH +
H+ for each triose.
 Hydrogen atom balance and charge balance are both maintained because the phosphate (Pi)
group actually exists in the form of a hydrogen phosphate anion (HPO42-), which dissociates to
contribute the extra H ion and gives a net charge of -3 on both sides.

Note that: Arsenate (AsO43−), an anion similar to inorganic phosphate may replace phosphate as a
substrate to form 1-arseno-3-phosphoglycerate. This, however, is unstable and readily hydrolyzes to form
3-phosphoglycerate, the intermediate in the next step of the pathway. As a consequence of bypassing this
step, the molecule of ATP generated from 1-3 bisphosphoglycerate in the next reaction will not be made,
even though the reaction proceeds. As a result, arsenate is an uncoupler of glycolysis during Arsenic
poisoning.

7. The transfer of the 1-phosphate group from 1,3-bis-phosphoglycerate to ADP by phosphoglycerate

kinase, which yields ATP and 3-phosphoglycerate.
 At this step, glycolysis has reached the break-even point: 2 molecules of ATP were consumed, and
2 new molecules have now been synthesized. This step, one of the two substrate-level
phosphorylation steps, requires ADP; thus, when the cell has plenty of ATP (and little ADP), this
reaction does not occur. Because ATP decays relatively quickly when it is not metabolized, this is
an important regulatory point in the glycolytic pathway.
 ADP actually exists as ADPMg−, and ATP as ATPMg2−, balancing the charges at -5 both sides.
8. The isomerization of 3-phosphoglycerate to 2-phosphoglycerate by phosphoglycerate
mutase(PGAM),

Note that: PGAM plays an important role in coordinating glycolysis and anabolic activity to promote cancer
cell proliferation. It is suggested that even under aerobic conditions cancer cells can show a high rate of
glycolysis and increased production of lactate, generating ATP faster than normal cells. However, to obtain
sufficient ATP and carbons to generate biomass [nucleotides, amino acids, lipids and nicotinamide adenine
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dinucleotide phosphate (NADPH)] cancer cells must uptake more glucose than normal cells to meet the
metabolic requirements of rapid cell proliferation owing to the low efficiency of glycolysis.

9. The dehydration of 2-phosphoglycerate to phosphoenolpyruvate by enolase.

10. The transfer of the phosphate group from phosphoenolpyruvate to ADP by pyruvate kinase, to yield
a second molecule of ATP.
A final substrate-level phosphorylation now forms a molecule of pyruvate and a molecule of ATP by
means of the enzyme pyruvate kinase. This serves as an additional regulatory step, similar to the
phosphoglycerate kinase step.

Most, but not all reactions in glycolysis are reversible; this can be indicated by
different as indicated in Fig arrows (.). Because it contains several irreversible reactions, the
pathway as a whole is also irreversible. However, alternate routes exist that bypass the irreversible
reactions and allow glucose to be synthesized from pyruvate.

Biochemical logic for the presence of regulatory steps

The existence of more than one point of regulation indicates that intermediates between those points enter
and leave the glycolysis pathway by other processes. For example, in the first regulated step, hexokinase
converts glucose into glucose-6-phosphate. Instead of continuing through the glycolysis pathway, this
intermediate can be converted into glucose storage molecules, such as glycogen or starch. The reverse
reaction, breaking down, e.g., glycogen, produces mainly glucose-6-phosphate; very little free glucose is
formed in the reaction. The glucose-6-phosphate so produced can enter glycolysis after the first control
point.

In the second regulated step (the third step of glycolysis), phosphofructokinase converts fructose-6-
phosphate into fructose-1,6-bisphosphate, which then is converted into glyceraldehyde-3-phosphate and
dihydroxyacetone phosphate. The dihydroxyacetone phosphate can be removed from glycolysis by
conversion into glycerol-3-phosphate, which can be used to form triglycerides. Conversely, triglycerides can
be broken down into fatty acids and glycerol; the latter, in turn, can be converted into dihydroxyacetone
phosphate, which can enter glycolysis after the second control point.
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Fig: 4.1. Glycolysis steps

Activity
1. Relate regulation of the rate limiting enzymes to homeostasis and biosynthesis?
2. Name and describe diseases related to failure in one or more of the steps of glycolysis?
3. Write the overall reactions of glycolysis.
4. Which of the following are products of cellular respiration?
A. Glucose; B. Pyruvate; C. Carbon dioxide; D. Acetyl CoA E.all except A

4.3.1.2. TCA cycle and ETC: Aerobic respiration

One fate of pyruvate is that it enters to TCA cycle for complete oxidation. But there are intermediate
processes that convert pyruvate to an acetyl coA. The enzyme complex converts pyruvate into Acetyl-CoA
by the following chemical changes:
 Decarboxylation of pyruvate (loss of CO2)
 Formation of acetyl group
 Linkage of acetyl group to coenzyme A forming acetyl - CoA.
The Tricarboxylic Acid (TCA) Cycle (Phase III)
The TCA cycle also called Krebs or Citric acid cycle, is considered as central pathway of aerobic
metabolism, as it serves two purposes-bioenergetics and biosynthesis:

1. Bioenergetics - The cycle carries out complex degradation of acetyl group in acetyl - CoA to CO2,
resulting in release of energy (ATP or GTP) and reducing power (NADH and FADH2).

TCA oxidizes two-carbon units, producing two molecules of CO2, one molecule of GP, and high-
energy electrons in the form of NADH and FADH2.

Steps of the Krebs cycle

Prior to the Krebs cycle, pyruvate first reacts with coenzyme A (CoA) to form a 2-carbon intermediate called
acetyl CoA. At the same time, carbon dioxide is released and NAD is converted to NADH. Acetyl CoA then
moves to the mitochondrial matrix. The reaction results in the production of two carbon dioxide molecules
and two NADH.
 The Krebs cycle begins with acetyl CoA combining with a 4-carbon compound to form a 6-
carbon compound known as citric acid.
 Citric acid is then broken down in the next series of steps, releasing two molecules of carbon
dioxide and generating one ATP, three NADH, and one FADH. FAD is another electron carrier
similar to NADH and NADPH.
 Finally, acetyl CoA and citric acid are generated and the cycle continues.
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Fig: 4.2. The TCA cycle

Recall that two molecules of pyruvate are formed during glycolysis, resulting in two “turns” of the Krebs
cycle for each glucose molecule. The net yield from the Krebs cycle is six carbon dioxide molecules, two
ATP, eight NADH, and two FADH2. NADH and FADH2 move on to play a significant role in the next stage of
aerobic respiration.

2. Biosynthesis - It supplies precursors for several biosynthetic pathways of amino acids, pyrimidines,
purines etc.
As already mentioned, the TCA cycle is also an important source of biosynthetic precursors
 e.g. α-ketoglutarate and oxaloacetate are used for synthesis of a number of amino
acids like glutamic acid, aspartic acid etc.
 Succinyl - CoA is used to form porphyrin ring of cytochromes, chlorophyll etc.
 Oxaloacetate can also be converted to phosphoenolpyruvate, which is a precursor of
glucose.
 Acetyl - CoA is the starting material for fatty acid biosynthesis.

4.3.1.3. Electron Transport Chain

In aerobic respiration, electron transport is the final step in the break-down of glucose. It also is the point at
which most of the ATP is produced. High-energy electrons and hydrogen ions from NADH and FADH2
produced in the Krebs cycle are used to convert ADP to ATP.
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Fig:4.3. Electron Transport Chain

As shown in Fig 4.3., electrons move along the mitochondrial membrane from one protein to another. As
NADH and FADH2 electrons, the energy carriers are converted to NADH release and FAD, and H ions are
released into the mitochondrial matrix. The H ions are pumped into the mitochondrial matrix across the
inner mitochondrial membrane. H ions then diffuse down their concentration gradient back across the
membrane and into the matrix through ATP synthase molecules in chemiosmosis. Electron transport and
chemiosmosis in cellular respiration are similar to these processes in photosynthesis. Oxygen is the final
electron acceptor in the electron transport system in cellular respiration. Protons and electrons are
transferred to oxygen to form water.
Activity
1.What is the overall chemical reaction of aerobic reparation?
2. How many ATP molecules are collected during electron transport chain and chemiosmosis from a single
glucose?

Biosynthesis
Biosynthesis is a multi-step, enzyme-catalyzed process where substrates are converted into more complex
products in living organisms. In biosynthesis, simple compounds are modified, converted into other
compounds, or joined together to form macromolecules. This process often consists of metabolic pathways.
Some of these biosynthetic pathways are located within a single cellular organelle, while others involve
enzymes that are located within multiple cellular organelles. Examples of these biosynthetic pathways
include the production of lipid membrane components and nucleotides. Biosynthesis is usually synonymous
with anabolism.

The prerequisite elements for biosynthesis include: precursor compounds, chemical energy (e.g. ATP), and
catalytic enzymes which may require coenzymes (e.g.NADH, NADPH). These elements create monomers,
the building blocks for macromolecules. Some important biological macromolecules include: proteins, which
are composed of amino acid monomers joined via peptide bonds, and DNA molecules, which are
composed of nucleotides joined via phosphodiester bonds.

Requirements of Biosynthesis
Sometimes all that is required for biosynthesis is for two substances to physically join together to make a
new physical substance, which is called a macromolecule.
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A very simplified equation for biosynthesis of one form of macromolecule: two substances join together to
form a new, more complex substance.

Photosynthesis
Self-test
1. Define photosynthesis.
2. What is an ultimate source of energy for all life forms?

Energy is transformed all around us every day. Batteries convert chemical energy into electric energy, and
radios convert electric energy into the energy carried by sound waves. Similarly, some autotrophs convert
light energy into chemical energy through photosynthesis.

The importance of photosynthesis

The processes of all organisms, from bacteria to humans require energy. To get this energy, many
organisms access stored energy by eating food. Carnivores eat other animals and herbivores eat plants.
But where does the stored energy in food originate? All of this energy can be traced back to the process of
photosynthesis and light energy from the sun. Photosynthesis is essential to all life on earth. It is the only
biological process that captures energy from outer space (sunlight) and converts it into chemical energy in
the form of Glyceraldehyde3-phosphate (G3P), which in turn can be made into sugars and other organic
compounds such as proteins, lipids, and nucleic acids. Plants use these compounds in all of their metabolic
processes; plants do not need to consume other organisms for food because they build all the molecules
they need. Unlike plants, animals need to consume other organisms to consume the molecules they need
for their metabolic processes.

The process of photosynthesis

During photosynthesis, molecules in leaves capture sunlight and energize electrons, which are then stored
in the covalent bonds of carbohydrate molecules. That energy within those covalent bonds will be released
when they are broken during cell respiration. How long lasting and stable are those covalent bonds? The
energy extracted today by the burning of coal and petroleum products represents sunlight energy captured
and stored by photosynthesis almost 200 million years ago.

Plants, algae, and some bacteria such as cyanobacteria are the only organisms capable of performing
photosynthesis. Because they use light to manufacture their own food, they are called photoautotrophs
(“self-feeders using light”). Other organisms, such as animals, fungi, and most other bacteria, are termed
heterotrophs (“other feeders”) because they must rely on the sugars produced by photosynthetic organisms
for their energy needs. A third very interesting group of bacteria synthesize sugars, not by using sunlight‟s
energy, but by extracting energy from inorganic chemical compounds; hence, they are referred to as
chemoautotrophs.
The importance of photosynthesis is not just that it can capture sunlight‟s energy. A lizard sunning itself on
a cold day can use the sun‟s energy to warm up. Photosynthesis is vital because it evolved as a way to
store the energy in solar radiation (the‟ „photo'' part) as high-energy electrons in the carbon-carbon bonds of
carbohydrate molecules (the ''synthesis'' part). Those carbohydrates are the energy source that
heterotrophs use to power the synthesis of
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ATP via respiration. Therefore, photosynthesis powers 99 percent of Earth‟s ecosystems. When a top
predator, such as a wolf, preys on a deer, the wolf is at the end of an energy path that went from nuclear
reactions on the surface of the sun, to light, to photosynthesis, to vegetation, to deer, and finally to wolf.

Other variant of photosynthesis

Commonly known photosynthetic processes is the one known as oxygenic photosynthesis. The other type
is termed as an oxygenic photosynthesis. The general principles of an oxygenic and oxygenic
photosynthesis are very similar, but oxygenic photosynthesis is the most common and is seen in plants,
algae and cyanobacteria.

During oxygenic photosynthesis, light energy transfers electrons from water (H2O) to carbon dioxide (CO2),
to produce carbohydrates. In this transfer, the CO2 is "reduced," or receives electrons, and the water
becomes "oxidized," or loses electrons. Ultimately, oxygen is produced along with carbohydrates.

On the other hand, an oxygenic photosynthesis uses electron donors other than water. The process
typically occurs in bacteria such as purple bacteria and green sulfur bacteria, which are primarily found in
various aquatic habitats.

An oxygenic photosynthesis does not produce oxygen. What is produced depends on the electron donor.
For example, many bacteria use the bad-eggs-smelling gas hydrogen sulfide, producing solid sulfur as a
byproduct.

Though both types of photosynthesis are complex, multistep affairs, the overall process can be neatly
summarized as a chemical equation.

Oxygenic photosynthesis is written as follows:

6CO2 + 12H2O + Light Energy → C6H12O6 + 6O2 + 6H2O

Similarly, the various an oxygenic photosynthesis reaction can be represented as a single generalized
formula:

CO2 + 2H2A + Light Energy → [CH2O] + 2A + H2O

The letter A in the equation is a variable and H2A represents the potential electron donor. For example, a
may represent sulfur in the electron donor hydrogen sulfide (H2S).

The photosynthetic apparatus

Self-test
1. What cellular components are essential to photosynthesis?
2. why plant leaves vary in color?

Plastids
Photosynthetic eukaryotic organisms contain organelles called plastids in their cytoplasm. Plastids
generally contain pigments or can store nutrients. Colorless and no pigmented leucoplasts store
fats and starch, while chloroplasts contain carotenoids and chloroplasts contain chlorophyll.
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Photosynthesis occurs in the chloroplasts; specifically, in the grana and stroma regions. The grana are the
innermost portion of the organelle; a collection of disc-shaped membranes, stacked into columns like
plates. The individual discs are called thylakoids. It is here that the transfer of electrons takes place. The
empty spaces between columns of grana constitute the stroma.

Chloroplasts are similar to mitochondria, the energy centers of cells, in that they have their own genome, or
collection of genes, contained within circular DNA. These genes encode proteins essential to the organelle
and to photosynthesis. Like mitochondria, chloroplasts are also thought to have originated from primitive
bacterial cells through the process of endosymbiosis.

Fig: 4.4. structure of chloroplast

Pigments
Pigments are molecules that bestow color on plants, algae and bacteria, but they are also responsible for
effectively trapping sunlight. Pigments of different colors absorb different wavelengths of light. Below are
the three main groups.

• Chlorophylls: These green-colored pigments are capable of trapping blue and red light.
Chlorophylls have three subtypes, dubbed chlorophyll a, chlorophyll b and chlorophyll c. There is
also a bacterial variant aptly named bacteriochlorophyll, which absorbs infrared light. This pigment
is mainly seen in purple and green bacteria, which perform an oxygenic photosynthesis.
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Fig: 4.5. basic structure of major photosynthetic pigments

• Carotenoids: These red, orange or yellow-colored pigments absorb bluish-green light. Examples
of carotenoids are xanthophyll (yellow) and carotene (orange) from which carrots get their color.
• Phycobilins: These red or blue pigments absorb wavelengths of light that are not as well absorbed
by chlorophylls and carotenoids. They are seen in cyanobacteria and red algae.

Antennae
Pigment molecules are associated with proteins, which allow them the flexibility to move toward light and
toward one another. A large collection of 100 to 5,000 pigment molecules constitutes antennae. These
structures effectively capture light energy from the sun, in the form of photons.
Ultimately, light energy must be transferred to a pigment-protein complex that can convert it to chemical
energy, in the form of electrons. In plants, for example, light energy is transferred to chlorophyll pigments.
The conversion to chemical energy is accomplished when a chlorophyll pigment expels an electron, which
can then move on to an appropriate recipient.

Reaction centers
The pigments and proteins, which convert light energy to chemical energy and begin the process of
electron transfer, are known as reaction centers.

The photosynthetic process

The reactions of plant photosynthesis are divided into those that require the presence of sunlight and those
that do not. Both types of reactions take place in chloroplasts: light dependent reactions in the thylakoid
and light-independent reactions in the stroma.
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Light-dependent reactions (also called light reactions): When a photon of light hits the reaction center, a
pigment molecule such as chlorophyll releases an electron.

How does this generate energy?

The released electron manages to escape by traveling through an electron transport chain, which
generates the energy needed to produce ATP (adenosine triphosphate, a source of chemical energy for
cells) and NADPH. The "electron hole" in the original chlorophyll pigment is filled by taking an electron from
water. As a result, oxygen is released into the atmosphere.

Light-independent reactions (also called dark reactions and known as the Calvin cycle): Light reactions
produce ATP and NADPH, which are the rich energy sources that drive dark reactions.

Although NADPH and ATP provide cells with large amounts of energy, these molecules are not stable
enough to store chemical energy for long periods of time. Thus, there is a second phase of photosynthesis
called the Calvin cycle in which energy is stored in organic molecules such as glucose.
Three chemical reaction steps make up the Calvin cycle: carbon fixation, reduction and regeneration.
These reactions use water and catalysts. The carbon atoms from carbon dioxide are ''fixed,'' when they are
built into organic molecules that ultimately form three-carbon sugars. These sugars are then used to make
glucose or are recycled to initiate the Calvin cycle again.

 In the first step of the Calvin cycle called carbon fixation, six carbon dioxide (CO) molecules
combine with six 5-carbon compounds to form twelve 3-carbon molecules called 3-
phosphoglycerat (3-PGA). The joining of carbon dioxide with other organic molecules is called
carbon fixation.
 In the second step, the chemical energy stored in ATP and NADPH is transferred to the 3-PGA
molecules to form high-energy molecules called glyceraldehyde 3phosphates (G3P). ATP
supplies the phosphate groups for forming G3P molecules, while NADPH supplies hydrogen
ions and electrons.
 In the third step, two G3P molecules leave the cycle to be used for the production of glucose
and other organic compounds. In the final step of the Calvin cycle, an enzyme called rubisco
converts the remaining ten G3P molecules into 5-carbon molecules called ribulose 1,5-
bisphosphates (RuBP). These molecules combine with new carbon dioxide molecules to
continue the cycle.
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Fig:4.6. Calvin cycle

Because rubisco converts inorganic carbon dioxide molecules into organic molecules that can be used by
the cell, it is considered one of the most important biological enzymes. Plants use the sugars formed during
the Calvin cycle both as a source of energy and as building blocks for complex carbohydrates, including
cellulose, which provides structural support for the plant.

Alternative Pathways
The environment in which an organism lives can impact the organism‟s ability to carry out photosynthesis.
Environments in which the amount of water or carbon dioxide available is insufficient can decrease the
ability of a photosynthetic organism to convert light energy into chemical energy. For example, plants in hot,
dry environments are subject to excessive water loss that can lead to decreased photosynthesis. Many
plants in extreme climates have altered native photosynthesis pathways to maximize energy conversion.

C4 plants one adaptive pathway that helps plants maintain photosynthesis While minimizing water loss is
called the C4 pathway. The C4 pathway occurs in plants such as sugar cane and corn. These plants are
called C4 plants because they fix carbon dioxide into four-carbon compounds instead of three-carbon
molecules during the Calvin cycle. C4 plants also have significant structural modifications in the
arrangement of cells in the leaves. In general, C4 plants keep their stomata (plant cell pores) closed during
hot days, while the four carbon compounds are transferred to special cells where CO 2 enters the Calvin
cycle. This allows for sufficient carbon dioxide uptake, while simultaneously minimizing water loss.

CAM plants Another adaptive pathway used by some plants to maximize photosynthetic activity is called
crassulacean acid metabolism (CAM photosynthesis). The CAM pathway occurs in water conserving plants
that live in deserts, salt marshes, and other environments where access to water is limited. CAM plants,
such as cacti, orchids, and the pineapple allow carbon dioxide to enter The leaves only at night, when the
atmosphere is cooler and more humid. At night, these plants fix carbon dioxide into organic compounds.
During the day, carbon dioxide is released from these compounds and enters the Calvin cycle. This
pathway Also allows for sufficient carbon dioxide uptake, while minimizing water loss.