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Bioenergetics

• Metabolism is a highly coordinated cellular activity


• By different chemical reactions, we obtain chemical energy in form of ATP
• In metabolic processes, convert nutrient molecules into the cell’s own
characteristic molecules and degrade biomolecules.
Bioenergetics describes the transfer and utilization of energy in biologic
systems
Examples of energy transfer within the human body
Muscle contraction: chemical energy to mechanical energy
Vitamin D formation: light energy to chemical energy
Photosynthesis: light energy to chemical energy in plants
Principles of Bioenergetics
• Its can enable to understand why these energy transfers occur.
• Biological energy transductions obey the laws of thermodynamics
• Energy cannot be created or destroyed, but can be changed from one form
to another
For any physical or chemical change, the total amount of energy in the
universe remains constant; energy may changed from or it may be
transported from one region to another
• Energy transfer will always proceed in the direction of increased entropy,
and the release of “free energy
In all natural processes the entropy of the universe increases
Free energy changes (ΔG)
 “Free energy” is referred to as Gibb’s free energy during energy transfers
 change in Gibb’s free energy is expressed as a ΔG
ΔG= ΔH - TΔS
ΔG= Change in Gibbs free energy of the reacting system (Gproducts– Greactives)
ΔH= Change in enthalpy of the reacting system(Hproducts – Hreactives), change in
heat content, heat release or absorbed
T= Absolute temperature
ΔS= Change in entropy of the reacting system(Sproducts – Sreactives), measure of
randomness
Sign of ΔG predicts the direction of a reaction
 (ΔG) provide a measure of the energetic feasibility of a chemical reaction
and therefore, allow prediction of whether a reaction can take place.
• Under the constant temperature and pressure changes in free energy,
determined by the degree to which two factors change during the
reaction, enthalpy (ΔH change in heat content of the reactants and
products) and entropy (ΔS, a measure of the change in randomness or
disorder of reactants and products
 When a reaction proceeds with the release of free energy, ΔG has a
negative value and the reaction is said to be exergonic
 In endergonic reactions, the system gains free energy and ΔG is positive
 If ΔG = 0, the reactants are in equilibrium
Change in free energy (ΔG)
ΔG of the forward and back reactions
• free energy of the forward reaction (A → B) is equal in magnitude but
opposite in sign to that of the back reaction (B → A).
• if ΔG of the forward reaction is −5 kcal/mol, then that of the back reaction is
+5 kcal/mol
standard free energy change ΔGo
• The change in free energy is represented in two ways, ΔG and ΔGo. The
• ΔG represents the change in free energy thus, the direction of a reaction at
any specified concentration of products and reactants.
• standard free energy change, ΔGo energy change when reactants and
products are at a concentration of 1 mol/L. pH = 7
• ΔGo can readily be determined from measurement of the equilibrium constant
ΔG depends on the concentration of reactants and products
ΔG of the reaction A → B depends on the concentration of the reactant and
product At constant temperature and pressure, the following relationship can
be derived
ΔG = ΔGo + RT ln[B]/[A]
ΔGo standard free energy change
(energy change when reactants and products are at a concentration of 1 mol/L)
R is the gas constant (1.987 cal/mol . degree)
T is the absolute temperature
[A] and [B] are the actual concentrations of the reactant and product
ln represents the natural logarithm
ΔG0
• ΔG'0 is the difference between the free energy content of the
products and the free energy contents of the reactants under
standard conditions
• ΔG0 = ΔG0 products– ΔG0 reactives
• When ΔG0 is negative, the products contain less free energy than
the reactants and the reaction will proceed spontaneously under
standard conditions
• When ΔG0 is positive, the products contain more free energy than the
reactants and the reaction will tend to go in the reverse direction
under standard conditions
ΔG to be negative
• consider the reaction:
Glucose 6-phosphate →fructose 6-phosphate
• concentration of reactant, glucose 6-phosphate, is high compared with
the concentration of product, fructose 6-phosphate.
• Ratio of the product to reactant is small, and RT ln([fructose 6-
phosphate] ⁄ [glucose 6-phosphate]) is large and negative, causing ΔG
to be negative. Thus, the reaction can proceed the forward direction.
Standard free energy change, ΔGo
• Τhe standard free energy change, ΔGo is equal to the free energy change, ΔG, under
standard conditions— when reactants and products are at 1 mol/L concentrations
• Under these conditions, the natural logarithm of the ratio of products to reactants is
zero (ln1 = 0) and, therefore, the equation becomes: ΔG = ΔGo + 0
ΔGo is predictive only under standard conditions: ΔGo can be used to predict the
direction a reaction under these conditions, ΔGo is equal to ΔG
Relationship between ΔGo and Keq: In a reaction A → B, a point of equilibrium is
reached at which no further net chemical change takes place—that is, when A is being
converted to B as fast as B is being converted to A. In this state, the ratio of [B] to [A] is
constant, regardless of the actual concentrations of the two compounds:
Keq =[B]eq/[A]eq
Keq is the equilibrium constant
ΔGo and Keq
• If the reaction is allowed to go to equilibrium at constant temperature
and pressure, then at equilibrium the overall free energy change (ΔG) is
zero.
• ΔG = 0 = ΔGo + RT ln[B]eq/[A]eq

• Actual concentrations of A and B are equal to the equilibrium


concentrations of reactant and product [A]eq and [B]eq, and their ratio as
shown above is equal to the Keq. Thus
• ΔGo = -RT ln Keq
ΔGo of two consecutive reactions are additive
• The standard free energy changes (ΔGo) are additive in any sequence of
consecutive reactions, as are the free energy changes (ΔG). For
example:
Glucose + ATP → glucose 6-P + ADP ΔGo = –4,000 cal/mol
Glucose 6-P → fructose 6-P ΔGo = +400 cal/mol
Glucose + ATP → fructose 6-P + ADP ΔGo = –3,600 cal/mol
ΔGs of a pathway are additive
• This additive property of free energy changes is very important in
biochemical pathways through which substrates must pass in a
particular direction (for example, A → B → C → D → ...).
• As long as the sum of the ΔGs of the individual reactions is negative,
the pathway can potentially proceed , even if some of the individual
reactions of the pathway have a positive ΔG.
• The actual rate of the reactions does,of course, depend on the
lowering of activation energies by the enzymes that catalyze the
reactions
ATP AS AN ENERGY CARRIER
This principle of bioenergetics explains how a thermodynamically
unfavorable (endergonic) reaction can be driven in the forward direction by
coupling it to a highly exergonic reaction through a common intermediate
• Reactions or processes that have a large positive ΔG, such as moving ions
against a concentration gradient across a cell membrane, are made
possible by coupling the endergonic movement of ions with a second,
spontaneous process with a large negative ΔG, such as the exergonic
hydrolysis of adenosine triphosphate (ATP).
• In the absence of enzymes, ATP is a stable molecule because its hydrolysis
has a high activation energy.
• Energy coupling in biologic reactions occurs when the energy-requiring
and the energy yielding reactions share a common intermediate.
Reactions are coupled through common intermediates
• Two chemical reactions have a common intermediate when they occur
sequentially
A+B→C+D
D+X→Y+Z
• D is the common intermediate and can serve as a carrier of chemical
energy between the two reactions.
• Many coupled reactions use ATP to generate a common intermediate.
These reactions may involve the transfer of a phosphate group from ATP
to another molecule.
• Other reactions involve the transfer of phosphate from an energy-rich
intermediate to adenosine diphosphate (ADP), forming ATP.
Energy carried by ATP
• ATP consists of a molecule of adenosine (adenine + ribose) to which three
phosphate groups
• If one phosphate is removed, ADP is produced; if two phosphates are
removed, adenosine monophosphate (AMP) results. The standard free
energy of hydrolysis of ATP,
• ΔGo, is approximately –7.3 kcal/mol for each of the two terminal
phosphate groups. Because of this large negative ΔGo, ATP is called a high-
energy phosphate compound
Compounds have large free energy change
• Phosphorylated compounds Thioesters (Acetyl-CoA)
• Phosphoenolpyruvate, 1,3-bisphosphoglycerate
• Phosphocreatine, ADP, ATP, AMP
• PPi, Glucose 1-phosphate, Fructose 6-phosphate
Thioesters
• In thioesters a sulfur atom is replaced the
usual oxygen in the ester bond
• Thioesters have large, negative standard
free energy change of hydrolysis.
• Acetyl coenzyme A is one of many
thioesters important in metabolism. The
acyl group in these compounds is activated
for trans-acylation, condensation or
oxidation-reduction reactions.
ELECTRON TRANSPORT CHAIN
 Energy-rich molecules, such as glucose, are metabolized by a series of
oxidation reactions ultimately yielding CO2 and water
 The metabolic intermediates of these reactions donate electrons to specific
coenzymes—nicotinamide adenine dinucleotide (NAD+) and flavin adenine
dinucleotide (FAD)—to form the energy-rich reduced coenzymes, NADH and
FADH2.
 These reduced coenzymes donate a pair of electrons to a specialized set of
electron carriers called the electron transport chain
 As electrons are passed down the electron transport chain, they lose much of
their free energy.
 This energy can be stored by the production of ATP from ADP and inorganic
phosphate (Pi). This process is called oxidative phosphorylation
Mitochondrion
• The electron transport chain is present in the inner
mitochondrial membrane and is the final common pathway by
which electrons derived from different fuels of the body flow to
oxygen.
• Electron transport and ATP synthesis by oxidative
phosphorylation proceed continuously in all tissues that contain
mitochondria.
• Outer membrane is permeable whereas Inner mitochondrial
membrane is impermeable to most ions and metabolites and
specialized carriers are required to move ions or molecules
across this membrane.
• Inner mitochondrial membrane is highly convoluted which
increase the surface area of the membrane
Matrix of the mitochondrion
 Gel-like solution in the interior of mitochondria is 50% protein
 Enzymes responsible for the oxidation of pyruvate, amino acids, fatty acids
(by β-oxidation), and involve in TCA cycle.
 In addition, the matrix contains NAD+ and FAD (the oxidized forms of the
two coenzymes that are required as hydrogen acceptors) and ADP and Pi ,
which are used to produce ATP
Organization of electron transport chain
 The inner mitochondrial membrane have five separate protein complexes,
called Complexes I, II, III, IV, and V.
 Complexes I–IV are part of the electron transport chain
 Each complex accepts or donates electrons to relatively mobile electron
carriers, such as coenzyme Q and cytochrome c.
 Complex V catalyzes ATP synthesis and so is referred to as ATP synthase
Electron transport chain

NADH, produced from a variety of oxidative (catabolic)


processes, is the substrate for Complex I.
Succinate, an intermediate of the TCA cycle, is the substrate for
Complex II
Reactions of the electron transport chain
• With the exception of coenzyme Q, all members of this chain are
proteins. These may function as enzymes as is the case with the
dehydrogenases may contain iron as part of an iron–sulfur center
• In the cytochromes may be coordinated with a porphyrin ring
• They may contain copper as does the cytochrome a + a3 complex

Formation of NADH: NAD+ is reduced to NADH by dehydrogenases that


remove two hydrogen atoms from their substrate.
 Both electrons but only one proton (that is, a hydride ion, :H–) are
transferred to the NAD+, forming NADH plus a free proton, H+.
NADH dehydrogenase
 The free proton plus the hydride ion carried by NADH are next transferred
to NADH dehydrogenase, a protein complex (Complex I) embedded in the
inner mitochondrial membrane. Complex I has a tightly bound molecule of
FMN, a coenzyme structurally related to FAD
 FMN accepts the two hydrogen atoms (2e– + 2H+), becoming FMNH2. NADH
dehydrogenase also contains iron atoms paired with sulfur atoms to make
iron–sulfur centers, necessary for the transfer of the hydrogen atoms to the
next member of the chain, coenzyme Q (ubiquinone).
Coenzyme Q
Coenzyme Q (CoQ) is a quinone derivative with a long, hydrophobic
isoprenoid tail
CoQ is a mobile carrier and can accept hydrogen atoms both from
FMNH2, produced on NADH dehydrogenase (Complex I), and from
FADH2, produced on succinate dehydrogenase (Complex II),
glycerophosphate dehydrogenase and acyl CoA dehydrogenase
CoQ transfers electrons to Complex III. CoQ, then, links the
flavoproteins to the cytochromes
Cytochromes
 The remaining members of the electron transport chain are cytochromes.
 Each contains a heme group (a porphyrin ring plus iron)
 Cytochrome iron is reversibly converted from its ferric (Fe3+) to its ferrous
(Fe2+) form as a normal part of its function as a reversible carrier of electrons
 Electrons are passed along the chain from CoQ to cytochromes bc1 (Complex
III), c, and a + a3 (Complex IV)
Cytochrome a + a3
 This cytochrome complex is the only electron carrier in which the heme iron
has an available coordination site that can react directly with O2, and so also is
called cytochrome oxidase. At this site, the transported electrons, O2, and free
protons are brought together, and O2 is reduced to water
 Cytochrome oxidase contains copper atoms that are required for this complex
reaction to occur.
Site-specific inhibitors
 Site-specific inhibitors of electron transport have been
identified
 These compounds prevent the passage of electrons by binding
to a component of the chain, blocking the oxidation/reduction
reaction.
 Inhibition of electron transport inhibits ATP synthesis because
these processes are tightly couple
 Incomplete reduction of oxygen to water produces reactive
oxygen species (ROS), such as superoxide, hydrogen peroxide
and hydroxyl radicals.
 ROS damage DNA and proteins, and cause lipid peroxidation.
 Enzymes such as superoxide dismutase, catalase, and
glutathione peroxidase are cellular defenses against ROS
Release of free energy during electron transport
• Free energy is released as electrons are transferred along the electron
transport chain
• The electrons can be transferred as hydride ions (:H–) to NAD+, as hydrogen
atoms (•H) to FMN, coenzyme Q, and FAD, or as electrons (e– ) to
cytochromes.
Redox pairs: Oxidation of one compound is always accompanied by reduction of
a second substance
 oxidation of NADH to NAD+ accompanied by the reduction of FMN to FMNH2
 Oxidation-reduction reactions can be written as the sum of two separate
half-reactions, one an oxidation reaction and the other a reduction
reaction
 NAD+ and NADH form a redox pair, as do FMN and FMNH2
 Redox pairs differ in their tendency to lose electrons.
 This tendency is a characteristic of a particular redox pair, and can be
quantitatively specified by a constant, Eo (the standard reduction potential),
with units in volts.
Standard reduction potential (Eo)
 The more negative the Eo of a redox pair, the
greater the tendency of the reductant member of
that pair to lose electrons.
 The more positive the Eo, the greater the tendency
of the oxidant member of that pair to accept
electrons.
 Therefore, electrons flow from the pair with the
more negative Eo to that with the more positive Eo
ΔGo is related to ΔEο : The change in free energy is related directly to the
magnitude of the change in Eo:
ΔGo = – n F ΔEo
n = number of electrons transferred (1 for a cytochrome, 2 for NADH, FADH2,
and coenzyme Q)
F = Faraday constant (23.1 kcal/volt . mol)
ΔEo = Eo of the electron-accepting pair minus Eo of the electron-donating pair
ΔGo = change in the standard free energy
ΔGo of ATP
 The standard free energy for the phosphorylation of ADP to ATP is +7.3
kcal/mol.
 The transport of a pair of electrons from NADH to oxygen via the
electron transport chain produces 52.58 kcal.
 Therefore, more than sufficient energy is available to produce three
ATP from three ADP and three Pi (3 x 7.3 = 21.9 kcal/mol),
 The remaining calories are used for ancillary reactions such as Ca2+
transport into mitochondria or released as heat
 sometimes expressed as a P:O ratio (ATP made per O atom reduced) of
3:1.
 P:O for FADH2 is 2:1 because Complex I is bypassed.
OXIDATIVE PHOSPHORYLATION
 The transfer of electrons down the electron transport chain is
energetically favored because NADH is a strong electron donor and
molecular oxygen is an avid electron acceptor.
 However, the flow of electrons from NADH to oxygen does not directly
result in ATP synthesis.
 Chemiosmotic hypothesis explains how the free energy generated by the
transport of electrons by the electron transport chain is used to produce
ATP from ADP + Pi
 Proton pump: Electron transport is coupled to the phosphorylation of
ADP by the transport (“pumping”) of protons (H+) across the inner
mitochondrial membrane from the matrix to the intermembrane space at
Complexes I, III, and IV.
Electron transport chain coupled to the transport of protons
 This process creates an electrical gradient (with more positive charges
on the outside of the membrane than on the inside) and a pH gradient
(the outside of the membrane is at a lower pH than the inside
 The energy generated by this proton gradient is sufficient to drive ATP
synthesis.
 Thus, the proton gradient serves as the common intermediate that
couples oxidation to phosphorylation.
Electron transport chain shown coupled to the transport of
protons
ATP synthase: The enzyme complex ATP synthase (Complex V) synthesizes
ATP using the energy of the proton gradient generated by the electron
transport chain.
It is also called F1/Fo ATPase because the isolated enzyme can catalyze the
hydrolysis of ATP to ADP and Pi
 The chemiosmotic hypothesis proposes that after protons have been
pumped to the cytosolic side of the inner mitochondrial membrane,
they reenter the matrix by passing through a channel in the membrane-
spanning domain (Fo ) of Complex V, driving rotation of Fo and, at the
same time, dissipating the pH and electrical gradients.
 Fo rotation causes conformational changes in the extra-membranous F1
domain that allow it to bind ADP + Pi, phosphorylate ADP to ATP, and
release ATP.
Oligomycin: This drug binds to the Fo , domain of ATP synthase, closing
the H+ channel, preventing reentry of protons into the mitochondrial matrix,
and thus preventing phosphorylation of ADP to ATP.

Uncoupling proteins (UCP)


 UCPs occur in the inner mitochondrial membrane
 These carrier proteins create a “proton leak,” they allow protons to re-
enter the mitochondrial matrix without energy being captured as ATP
 The energy is released as heat, and the process is called nonshivering
thermogenesis.
 UCP1, also called thermogenin, is responsible for the heat production in
the brown adipocytes of mammals.
c. Synthetic uncouplers:
Electron transport and phosphorylation can also be uncoupled by
compounds that increase the permeability of the inner mitochondrial
membrane to protons.
2,4-dinitrophenol, a lipophilic proton carrier that readily diffuses through
the mitochondrial membrane.
This uncoupler causes electron transport to proceed at a rapid rate without
establishing a proton gradient, much as do the UCPs
Again, energy is released as heat rather than being used to synthesize ATP.
In high doses, aspirin and other salicylates uncouple oxidative phos -
phorylation.
This explains the fever that accompanies toxic overdoses of these drugs.
Membrane transport systems
The inner mitochondrial membrane is impermeable to most charged or
hydrophilic substances, numerous transport proteins that permit passage
of specific molecules from the cytosol to the mitochondrial matrix.
ATP-ADP transport: The inner mitochondrial membrane requires
specialized carriers to transport ADP and Pi from the cytosol (where ATP is
used and converted to ADP in many energy requiring reactions) into
mitochondria, where ATP can be resynthesized.
 An adenine nucleotide carrier imports one molecule of ADP from the
cytosol into mitochondria, while exporting one ATP from the matrix back
into the cytosol
 A phosphate carrier is responsible for transporting Pi from the cytosol
into mitochondria
Transport of reducing equivalents Inner mitochondrial membrane lacks an
NADH transporter, and produced in the cytosol cannot directly enter the
mitochondrial matrix. However, two electrons of NADH are transported from the
cytosol into the matrix using substrate shuttles.
 Glycero phosphate shuttle, two electrons are transfer from NADH to
dihydroxyacetone phosphate by cytosolic glycerophosphate dehydrogenase.
 The glycerol 3-phosphate produced is oxidized by the mitochondrial isozyme
as FAD is reduced to FADH2. CoQ of the ETC oxidizes FADH2.
 The glycero phosphate shuttle, therefore, results in the synthesis of two ATPs
for each cytosolic NADH oxidized.
 Malate-aspartate shuttle produces NADH in the mitochondrial matrix and,
therefore, yields three ATPs for each cytosolic NADH oxidized by malate
dehydrogenase as oxaloacetate is reduced to malate. A transport protein
carries malate into the matrix

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