Cell Bioenergetics

Energy production, ATP generation by

different processes, free energy,
Energy coupling:
 Energetics
• Energy is defined as the ability to do work. Some organisms use chemical energy.
Chemical energy is the energy released when organic or inorganic compounds are
oxidized. Microorganisms trap energy from biochemical reactions. Many of these
microbial metabolic mechanisms are also used by higher organisms (including
humans) to obtain energy.

Energy Requirements of microbial cell:

 
A living cell requires energy to perform many different kinds of work, including:
 
• Construction of structural parts of the cell such as wall, membrane, or external
appendage.
• Synthesis of enzymes, nucleic acids, polysaccharides, phospholipids, and other
chemical components of the cell.
• Repair of damage and maintenance of the cell in good condition.
• Growth and multiplication.
• Accumulation of nutrients and excretion of waste products
• Motility.
 First law of thermodynamics:
• In any physical or chemical change, the total amount of energy in
the universe remains constant. In other words, energy is
conserved. The mathematical expression of the first law is,
• ∆E = EB-EA = Q-W.
 
• Here,
• EA = Energy of a system at the start of a process
• EB = Energy of a system at the end of a process
• Q = heat absorbed by the system
• W = Work done by the system
 
• An important feature of the first law is that the change in energy
of a system depends only on the initial and final states and not on
the path of the transformation.
 Second law of thermodynamics:

• All physical and chemical changes tend to proceed in such a

direction that useful form of energy undergoes irreversible
degradation into a randomized (useless form) called entropy.
•  
• There are two types of useful energy:
•  
• Free energy: The energy available to do work usually at a constant
temperature and pressure.
• Heat energy: The energy which can do work only through a
change of temperature.
•  
• Entropy is the energy in a state of randomness or disorder. It is
unavailable useless energy.
 Equation of free energy:
Free energy is denoted by the symbol G (or, F in older literature). In 1878, Josiah Willard Gibbs created
the free-energy function by combining the first and second law of thermodynamics. The basis
equation is,
 
• ∆G = ∆H - T∆S,

• Here, ∆G = Change in free energy of a system at constant pressure and temperature.

• ∆H = Change in enthalpy of the system
• ∆S = Change in entropy.

• The enthalpy change can be expressed by the equation

• ∆H = ∆E + P∆V

• Here, ∆E = Change in internal energy

• P = Pressure
• ∆V = Volume change.

• The volume change ∆V is small for nearly all biochemical reactions, and so, it is negligible.

• So, ∆H = ∆E + Px0
• ∆H = ∆E

• Therefore, ∆G = ∆E - T∆S
• Thus the ∆G of a reaction depends both on the change in
internal energy and on the change in entropy of the system.
The change in free energy (∆G) of a reaction, in contrast
with the change in internal energy (∆E) of a reaction, is a
valuable criterion of whether it can occur spontaneously;
 
• A reaction can occur spontaneously only if ∆G is negative.
• A system is at equilibrium and no net change can take place
if ∆G is zero.
• A reaction cannot occur spontaneously if ∆G is positive. An
input of free energy is required to drive such a reaction.
• The ∆G of a reaction is independent of the path of the
transformation. The ∆G of a reaction depends only on the free
energy of the products (the final state) minus that of the reactants
(the initial state). The mechanism of a reaction has no effect on
∆G.
•  
• For example: The ∆G for the oxidation of glucose to CO2 and H2O is
the same whether it occurs by combustion in vitro or by a series of
many enzyme catalyzed steps (EMP, ED, HMP pathways) in cell.
•  
• ∆G provides no information about the rate of a reaction. A
negative ∆G indicates that a reaction can occur spontaneously, but
it does not signify that it will occur at a perceptible rate. The rate
of a reaction depends on the free energy of activation, which is
unrelated to ∆G.
 Energy Coupling:
• The dissimilation of nutrients and the synthesis of cell constituents
involve numerous chemical reactions, each catalyzed by a specific
enzyme.
• In course of any chemical reaction, the chemical energy is either
released or absorbed. A chemical reactions that liberates energy is
called an exergonic reaction, whereas a chemical reaction that
takes up energy is called an endergonic reaction.
• Exergonic reactions are associated with the dissimilation of a
nutrient or chemical substrate, while endergonic reactions are
associated with synthesis of cell constituents.
• In a living organism, the exergonic reactions provide the energy to
fuel the endergonic reactions. In order to link these reactions,
organisms have developed a process called energy coupling.  
The energy transfer compounds that are of most use to a cell are those capable of
transferring large amounts of energy (High energy- transfer compounds) occur in
cells. But, one is by far most important: Adenosine triphosphate (ATP).
 General role of ATP in energy coupling
• ATP is composed of one molecule of the purin base adenine, one
molecule of the pentose sugar ribose, and three phosphate groups.
• It is formed by adding a phosphate group to adenosine
diphosphate (ADP) which has only two phosphate groups:
Energy
• ADP + phosphate ATP + Water
• A large amount of energy is needed to form the chemical bond
linking the third phosphate group to ADP, for this reason, the bond
is called a high-energy phosphate bond. The energy trapped in the
high energy phosphate bond of ATP can be liberated if the bond is
later broken
• Just as money constitutes a common medium for buying and
selling of items in a society, so ATP constitutes the ‘energy
currency’ of a cell during the exchange of chemical energy between
many different kinds of exergonic and endergonic chemical
reactions.
The flow of chemical energy from dissimilation or nutrient molecules to ATP,
and then from ATP to the energy-requiring (endergonic) reactions of a
microbial cell.

Exergonic reactions
associated with
dissimilation
Nutrient End
molecules product
Energy

Some energy lost as heat

Chemical
energy

Energy coupling
ADP+Phosphate ATP
system

Endergonic reactions associated with

synthesis of cell constituents and other cell
functions that require energy
 Energy rich compounds/ High energy compounds:

• In living organisms, chemical energy released in oxidation-reduction reaction is most

commonly transferred to a variety of phosphate compounds in the form of high
energy phosphate bonds; these compounds then function as the energy source to
drive energy-requiring reactions in the cell or serves as intermediates in the
conversion of energy into useful work.
• They are called energy-rich or high energy compounds because they exhibit a large
decrease in free energy when they undergo hydrolytic reactions.
• They are in general unstable to acid, to alkali, and to heat. In phosphorylated
compounds, phosphate groups are attached via oxygen atoms by ester or anhydride
bonds.

• Examples: Phosphoenolpyruvate (PEP)

• 1,3-bisphosphoglycerate
• Acetyl phosphate
• ATP
• ADP
•  
• Not all phosphate bonds are high energy bonds. Some contain low energy phosphate
bonds. Cells usually do not use them directly as energy source, because of low ∆G°.
Example: AMP, Glucose-6-phosphate.
 ATP is universal currency of free energy in biological
system:
Out of all energy rich compounds, ATP is the universal energy currency in biological
systems:
•  
ATP is a nucleotide consisting of an adenine, a ribose and a triphosphate unit. ATP is an
energy-rich molecule because of its triphosphate unit contains two phospho-
anhydride bonds. A large amount of free energy is liberated when ATP is hydrolyzed
to ADP and orthophosphate (Pi) or when ATP is hydrolyzed to AMP and
pyrophosphate (Pi).
•  
• ATP + H2O ↔ ADP + Pi + H+ , ∆G° = -7.3 Kcal/mol
•  
• ATP + H2O ↔ AMP + PPi + H+ , ∆G° = -7.3 Kcal/mol
•  
ATP, AMP and ADP are inter-convertible. The enzyme adenylate kinase (also called
myokinase) catalyzes the reaction.
Adenylate kinase
• ATP + AMP ADP + ADP  
• The free energy liberated in the hydrolysis of anhydride bond of ATP is used to drive
reactions that require an input of free energy (endergonic), such as muscle
contraction. In turn, ATP is formed from ADP and Pi when fuel molecules are
oxidized in chemotrophs or when light is trapped by phototrophs. This ATP-ADP cycle
is the fundamental mode of energy exchange in biological systems.
Other energy rich compounds, analogous to ATP, are GTP, UTP, CTP. But they are not
used by the cell. Because their phosphorous dissociation tendency is lesser than ATP.
The diphosphate forms of GTP, UTP, CTP are GDP, UDP, CDP, respectively. Enzymes
catalyze the transfer of the terminal phosphoryl group from one nucleotide to
another as in the reactions,
•  
• ATP + GDP ↔ ADP + GTP, catalyzed by nucleoside diphosphokinase
•  
• ATP + GMP ↔ ADP + GDP, catalyzed by nucleoside monophosphokinase.
•  
• So, enzymes have specificities. But most enzyme use ATP as energy source, not CTP,
GTP or UTP.
•  
Again, the formation of GTP, GDP etc. require ATP.
 
• From all these point of views, ATP is the universal currency of energy.
 Generation of ATP/energy:
• High energy phosphate bonds trap energy and this energy is generated by a process
called phosphorylation. Phosphorylation is the addition of a phosphate group to a
compound. ATP is formed (or, energy is generated) by phosphorylation of ADP, with
energy for the addition provided by an exergonic reaction.
 
• Energy
• ATP
• ADP + Pi
•  
• There are three general ways in which phosphorylation of ADP can occur/ energy can
be generated/ ATP can be formed.
•  
Substrate-level phosphorylation: A process in which the phosphate group of a chemical
compound is removed and directly added to ADP. Example: Phosphoenolpyruvate
(PEP) contain high energy phosphate group. PEP can donate this phosphate to ADP
which then form ATP.
Oxidative phosphorylation: A process by which the energy liberated by chemical
oxidations of nutrients chemical compounds is used for the synthesis of ATP from
ADP.
Photophosphorylation: A process in which the energy of light is used for the synthesis of
ATP from ADP.
 Substrate –level phosphorylation:

 
• Sometimes when phosphate bond in a chemical compound is not highly
energetic to be dissociated, then the rearrangement of atoms within
chemical compounds may result in a new compound that contains a high
energy phosphate bond. Such rearrangements occur when cells break-
down nutrients into chemical compounds. The phosphate group involved
in this bond can then be transferred directly to ADP, forming ATP, which
now contains the high energy phosphate bond.
•  
• Example: In a cell glucose break down and one compound that may result
from glucose is 2-phosphoglyceric acid. It has phosphate group which
does not have sufficient energy to be dissociated. The cell then rearranges
the atoms in the 2-phosphoglyceric acid by removing a molecule of water,
thus forming a new compound, PEP. It has a high energy phosphate bond.
In fact the bond has sufficient energy to allow the phosphate group on
PEP to be transferred directly to ADP forming ATP.
 Oxidative phosphorylation
• All oxidation reactions liberate energy, and
many organisms have developed ways to use
the energy from chemical oxidations for ATP
synthesis.
• The overall process of using the energy from
oxidation reactions to make ATP from ADP is
called oxidative phosphorylation.
 The sequence of events in this process can
be summarized as follows:
• Energy is liberated by an integrated series of sequential
chemical oxidation reactions called an Electron Transport
System/chain (ETC).
 
• The energy is stored temporarily in the form of a Proton
Motive Force (PMF)
 
• The proton motive force powers the synthesis of ATP from
ADP.
 
• So, electron transport system and proton motive force
together govern the oxidative phosphorylation
 Electron transport system/chain (ETC):

• A cell uses an integrated series of sequential

oxidation reactions called an electron
transport system, which liberates the energy
in several smaller increments.
• Two functions of ETC:
– 1. to accept electrons from an electron donor and
transfer them to an electron acceptor;
– 2. to conserve some of the energy released during
electron transfer for the synthesis of ATP.
• Process of sequential oxidation reaction: An
electron transport system is composed of a
series of O/R systems in which successive O/R
system has a greater ability to gain electrons
(a greater oxidizing ability) than the one
preceding it.
• The following is a schematic illustration of an
ETC
Mitochondrial Electron transport chain
• Complex I: NADH
dehydrogenase or NADH
oxidoreductase

• CompleQ/Coenzyme Q
(Ubiquinonx II: Succinate
reductase
• Complex-III: cytochrome C
oxidorereductase
• Complex IV: Cytochrome C
oxidase
• e)
• Cyt C: Cytochrome C
• The system begins with an electron-donor, a
reduced compound (X(red)) which provides
the electrons. This reduced compound is
either a nutrient that has been taken in by the
cell or a compound resulting from the
breakdown of a nutrient. For instance; some
microorganisms use lactic acid as an electron
donor;
Lactic Pyruvic acid + 2H+ + 2e-
acid
• The electrons from the electron donor are
removed by an initial O/R system. This O/R system
is oxidized by the next O/R system, again and
again and so forth. Finally, the electrons are taken
up by a terminal electron acceptor, an oxidizing
compound (Y(ox)) obtained from the cells
environment. For instance, aerobic organisms use
oxygen as the terminal electron acceptor; after
accepting the electrons from the last O/R system,
the oxygen becomes reduced to water.

½ O2 + 2e- H2O
+ 2H+
• Anaerobic organism uses a chemical such as
nitrate, sulfate, or fumaric acid as terminal
electron acceptor. The importance of an ETC lies
in the fact that energy is liberated at each step in
the sequential series of oxidations. At some steps,
the amount of energy liberated is so great that it
can allow ATP to make.
•  
• In a bacterium, ETC is located in the cytoplasmic
membrane, whereas in a eukaryotic cell it is in the
inner membrane of mitochondria
 Proton Motive Force (PMF):
• In 1978, the biochemist Peter Mitchell received a Nobel Prize for
discovering the way by which energy liberated by an electron transport
system is used for the synthesis of ATP (Chemiosmotic theory).
• The energy liberated in an electron-transport system is used to pump
protons (H+) across the membrane.
• Some of these protons are derived from the hydrogen atoms of the
electron donor; others are the hydrogen ions that occur in water.
• After the protons are pumped across the membrane, they cannot easily
return, because the membrane is not permeable to protons.
• Therefore, the continued operation of an electron transport system results
in accumulation of protons on one side of the membrane ( outside the
bacterial cell) and a deficit of protons on the opposite side (inside the
bacterial cell).
• The net result is that one side of the membrane becomes much more
positively charged and acidic (low pH) than the other side. So, a pH
gradient occurs.
• The universal distribution of protons (pH gradient) and electric charges
(electron potential) across the membrane represents an important form of
potential energy called the proton-motive force, which is used to
synthesize ATP.
• The PMF represents potential energy in much the same way that a
body of water held back by a dam represents potential energy. If a
gate in the dam were opened, the water would flow down from
the higher elevation toward a lower level; this water flow can
operate a turbine and generate hydroelectric power.
•  
• The membrane of a cell is like a dam- it separates protons at a
high concentration on one side from a lower portion
concentration on the other side. If a channel specific for protons is
present in the membrane, the protons will flow ‘downhill’ to the
side where they are less concentrated. This proton flow can be
used by a cell to perform work.
•  
• Certain specific channels do exist in the cytoplasmic membranes
that allow passage of protons back to the other side of the
membrane. These channels occur within the molecule of an
enzyme called adenosine triphosphatase (ATPase), which spans
the membrane.
 Formation of ATP:

• ATPase enzyme contains two main parts, a multisubunit headpiece

present in the inside of the membrane and a proton- conducting
tailpiece that spans the membrane. This enzyme catalyzes the
reversible reaction between ATP and ADP + Pi
•  
• Operating in one direction, this enzyme catalyzes the formation of
ATP by allowing the controlled reentry of protons across the
energized membrane. Just as the formation of proton gradient was
energy-driven, the controlled dissipation of the proton-motive
force is energy-releasing, and some of the energy is conserved in
the synthesis of ATP in a process called oxidative phosphorylation.
•  
• ATPase can also catalyze the reverse reaction, that is, the hydrolysis
of ATP and the extrusion of 4H+ to the outer portion of the
membrane. This results in the conversion of phosphate bond
energy to the energy of a proton-motive force
• The biochemical mechanisms of oxidative phosphorylation are
illustrated in the following figure:
 Photophosphorylation
• Photophosphorylation is the overall process in which light is used as a source of
energy for ATP synthesis. The general way in which photophosphorylation occurs is
as follows:
 
– Light is used to generate PMF
– The PMF then powers ATP synthesis.
 
• The most important example of photophosphorylation is the type carried out by
cyanobacteria, algae and green plants. The phosphorylation system in these
organisms occurs within the cells in special flattened membranous sacs called
thylakoids.
 
• In cyanobacteria (Prokaryotes), the thylakoids occur directly within the cytoplasms,
but in the cells of algae and green plants (eukaryotes), they are contained within the
green chloroplasts. Thylakoid membranes contain chlorophyll (chl), a light absorbing
green pigment that plays a major role in the photophosphorylation process.
• Cyanobacteria, algae and green plants are able to
use CO2 as their sole source of carbon; that is,
they are autotrophic organisms. They reduce the
CO2 to carbohydrate (CH2O)x by a process called
CO2 fixation. This process requires two
components: 
• 1. ATP, which serves as an energy source and is
made by photophosphorylation;
• 2. NADPH2, the reduced form of a coenzyme
called nicotinamide adenine dinucleotide
phosphate (NADP). The NADPH2 is used as an
electron donor for reduction of CO2.
Mechanism of photosynthetic electron flow:

• The generation of ATP, and NADPH2 depends on the activity of two

different kinds of chlorophyll-containing reaction centers, called
photosystem I (PS-I) and photosystem II (PS-II). They are located in
thylakoid membranes. The two photosystems work together where
three main steps are involved
•  
• 1. When light is absorbed by the chlorophyll molecules in PSI, the
energy of the light raises the molecules to an excited state, causing
an electron to be ejected from each. These ejected electrons are
used to reduce NADP to NADPH2. This leaves the chlorophyll of PSI
temporarily deficient in electrons, giving it a positive charge
2 Chl (PSI) NADP + 2H+

2e-

2 Chl (PSI)+ NADPH2

• Similarly light is absorbed by the chlorophyll of PSII and causes electrons
to be ejected by this photosystem. These electrons pass along an electron-
transport system and reach the electron deficient chl+ of PSI. This electron
transport system differs from oxidative electron-transport system in that,
the electron donor chl (PSII) and the terminal electron acceptor chl (PSI) +
are supplied by the cell itself, rather than being obtained from the
environment. Nevertheless, the result is the same as with oxidative
phosphorylation- a proton motive force is generated across the membrane
and is used to power the ATP synthesis.
•  
• At this point, Chl (PSII)+ is still deficient in electrons. However, Chl (PSII)+ is
a very strong oxidizing agent- so strong that it can regain electrons by
removing them from molecules of water. This oxidation of water results in
the formation of gaseous oxygen:

H2O 2Chl (PSII)+

2e-

½ O2 + 2H+ 2 Chl (PSII).

Cyclic Photophosphorylation
Noncyclic phototphosphorylation
Figure of photophosphorylation: (PMF and generation of ATP in photophosphorylation are as described in oxidative
phosphorylation)
Chemicals effecting the operation of ETC and creation of a PMF:

• Two classes of chemicals are known to effect the operation of ETC and
creation of a PMF. These are: 1. Inhibitors and 2. Uncouplers or uncoupling
agents.
•  
• Inhibitors: Inhibitors block electron flow and thus ATP synthesis.
Examples: a) Carbon monoxide (CO): It prevents the reduction of O2 to
H2O by preventing terminal electron acceptor to donate electron to O2. b)
Cyanide (CN-), Hydrogen sulfide (H2S), or azide (N3-). They bind tightly to
cytochromes (the electron carriers) and block electron transport.
•  
• Uncouplers: Uncouplers inhibit ATP synthesis without affecting the
ATPase. The PMF which is developed by ETC, is destroyed by them.
Examples: Dinitrophenol and dicumarol. These are lipid soluble substances
that increase membrane permeability, thereby promoting the leakage of
protons across the membrane. The latter results in dissipation of the
proton motive force and hence inhibition of ATP synthesis.