BIOCHEMISTRY 2

TOPIC 2:
PRINCIPLES OF BIOENERGETICS,
GLYCOLYSIS, GLUCONEOGENESIS,
PENTOSE PHOSPHATE PATHWAYS
Vũ Thanh Ngọc, PhD
Department of Applied Chemistry
School of Biotechnology
HCMC International University
vtngoc@hcmiu.edu.vn
Topic 2 Week
1. Principles of bioenergetics 2

2. Glycolysis 3

3. Gluconeogenesis 3

4. Pentose phosphate pathways 3

 2.1. PRINCIPLE OF BIOENERGETICS
1. Bioenergetics and thermodynamics
Laws of thermodynamics
Quantitative relationships among free energy, enthalpy, and entropy
2. Phosphoryl group transfers and ATP
Roles of ATP in biological energy exchange
3. Biological oxidation-reduction reactions
Energetics of electron-transfer reactions
Electron carriers: NAD and NADP, FAD and FMN
 WHAT IS BIOENERGETICS?

Bioenergetics is the quantitative study of:

 The energy transductions that occur in living cells of the nature

 Function of the chemical processes underlying these transductions

 WHAT IS BIOENERGETICS?

How does the cells tap into this process?

 TYPES OF ENERGY IN BIOLOGICAL SYSTEMS

Potential energy:
 Stationary
 Exist as chemical bonds, concentration gradients, and electrical
charges in the cells

Kinetic energy:
 Energy in motion
 Result in the movement of molecules
 BIOLOGICAL ENERGY TRANSFORMATIONS
OBEY THE LAWS OF THERMODYNAMICS

First law – energy conservation:

“For any physical or chemical change, the total amount of energy in
the universe remains constant; energy may change form or it may be
transported from one region to another, but it cannot be created or
destroyed.”
 ENERGY CONSERVATION
In thermodynamics, a system can be classified based on the exchange
of energy and materials with another system:
Isolated system: NO energy or material can be exchanged
with the surrounding
Closed system: energy but NO material can be exchanged
with the surroundings
Open system: BOTH material and energy can be exchanged
with the surroundings

 Living cells or organisms are open systems

 BIOLOGICAL ENERGY TRANSFORMATIONS
OBEY THE LAWS OF THERMODYNAMICS

Second law – entropy expansion:

“In all natural processes, the entropy of the universe increases.”

 Reactions occur spontaneously only when they increase disorder

in the system and its surroundings. Otherwise, the system will
REQUIRE energy (Gibbs free energy) for non-spontaneous reactions.
 RELATIONSHIP AMONG FREE ENERGY,
ENTHALPY AND ENTROPY
Gibbs free energy, G: amount of energy capable of doing work during
a reaction at constant temperature and pressure
 ΔG > 0, endergonic reactions (non-spontaneous)
ΔG < 0, exergonic reactions (spontaneous)
Enthalpy, H: heat content of the reacting system
 ΔH > 0, endothermic reactions
ΔH < 0, exothermic reactions
Entropy, S: the randomness or disorder in a system
 RELATIONSHIP AMONG FREE ENERGY,
ENTHALPY AND ENTROPY
Under constant temperature and pressure:

ΔG = ΔH – TΔS

J/mol or cal/mol K J/mol.K or cal/mol.K

1 cal = 4.184 J
K: Kelvin, 25 oC = 298 K
RELATIONSHIP BETWEEN EQUILIBRIUM CONSTANT
AND STANDARD FREE ENERGY CHANGE

equilibrium constant Keq:

[A], [B], [C], and [D]: molar concentrations of the reaction

components at the point of equilibrium.
 RELATIONSHIP BETWEEN EQUILIBRIUM CONSTANT
AND STANDARD FREE ENERGY CHANGE

ΔG’º = -RT ln K’eq

Gas constant, R = 8.315 J/mol.K = 1.987 cal/mol.K)

Standard condition: T = 298 K (25oC), 1 atm, [H+] = 1 M or pH = 0
Biochemical standard condition: pH = 7, concentration of water = 55.5 M
 ΔG’º and K’eq: standard transformed constants
characteristic for each reaction
RELATIONSHIP BETWEEN EQUILIBRIUM CONSTANT
AND STANDARD FREE ENERGY CHANGE

Small free energy change leads to

exponential change in equilibrium constant
RELATIONSHIP BETWEEN EQUILIBRIUM CONSTANT
AND STANDARD FREE ENERGY CHANGE
RELATIONSHIP BETWEEN EQUILIBRIUM CONSTANT
AND STANDARD FREE ENERGY CHANGE

Standard conditions,
NOT conditions existing in cells
 Actual free-energy change?
 ACTUAL FREE-ENERGY CHANGES DEPEND ON
REACTANT AND PRODUCT CONCENTRATIONS

 When ΔG < 0, the reaction is thermodynamically feasible, but may not

happen by itself.
 Ex: combustion of firewood to CO2 and H2O is very favorable
thermodynamically, but firewood remains stable for years. Why?
 the activation energy for the combustion reaction is higher than
the energy available at room temperature
 ACTUAL FREE-ENERGY CHANGES DEPEND ON
REACTANT AND PRODUCT CONCENTRATIONS

To occur, the reaction needs help to overcome the activation barrier

of the transition state.
STANDARD FREE-ENERGY CHANGES ARE ADDITIVE

2 sequential chemical reactions:

 THE ROLE OF ATP IN METABOLISM
ATP is the chemical link between catabolism and anabolism.
Heterotrophic cells obtain free energy (chemical energy)
via catabolism of nutrient molecules.
 Use that energy to make ATP from ADP and Pi
 Exergonic conversion of ATP to ADP and Pi, or to AMP
and PPi, is coupled to endergonic processes:

 Synthesis of macromolecules

 Membrane transport against concentration gradient

 Mechanical motion
THE FREE-ENERGY CHANGE FOR ATP
HYDROLYSIS IS LARGE AND NEGATIVE

Chemical basis:
1. ATP is unstable due to charge repulsion
2. The leaving inorganic Pi undergoes
resonance stabilization
3. ADP is immediately ionized to ADP3- + H+
OTHER PHOSPHORYLATED COMPOUNDS AND THIOESTERS
ALSO HAVE LARGE FREE ENERGIES OF HYDROLYSIS
Phosphoenolpyruvate (PEP)
OTHER PHOSPHORYLATED COMPOUNDS AND THIOESTERS
ALSO HAVE LARGE FREE ENERGIES OF HYDROLYSIS
1,3-bisphosphoglycerate
OTHER PHOSPHORYLATED COMPOUNDS AND THIOESTERS
ALSO HAVE LARGE FREE ENERGIES OF HYDROLYSIS
Phosphocreatine
OTHER PHOSPHORYLATED COMPOUNDS AND THIOESTERS
ALSO HAVE LARGE FREE ENERGIES OF HYDROLYSIS
Thioesters such as acetyl-CoA
OTHER PHOSPHORYLATED COMPOUNDS AND THIOESTERS
ALSO HAVE LARGE FREE ENERGIES OF HYDROLYSIS
OTHER PHOSPHORYLATED COMPOUNDS AND THIOESTERS
ALSO HAVE LARGE FREE ENERGIES OF HYDROLYSIS

To summarize, for hydrolysis reactions with large, negative ΔG’º,

the products are more stable than the reactants because:
(1) the bond strain in reactants due to electrostatic repulsion is relieved
by charge separation
(2) the products are stabilized by ionization

(3) the products are stabilized by isomerization (tautomerization)

(4) the products are stabilized by resonance

 ATP PROVIDES ENERGY BY GROUP TRANSFERS,
NOT BY SIMPLE HYDROLYSIS

Besides direct hydrolysis, energy of ATP

breakdown is coupled to endergonic
transformations of substrates by transferring a
phosphoryl, pyrophosphoryl, or adenylyl group
from ATP to a substrate or enzyme molecule

ATP hydrolysis in two steps

THE ENERGY RANK OF SOME BIOLOGICAL
PHOSPHORYLATED COMPOUNDS
 THE ROLE OF INORGANIC POLYPHOSPHATE

 Inorganic polyphosphate (polyP) is a linear polymer composed of many Pi

residues linked through phosphoanhydride bonds.
 Present in all organisms

 May serve as a reservoir of phosphoryl groups with high group transfer

potential.
The transfer of phosphoryl group is not the only way of channeling
energy inside the cell. Another way is electron transfer in oxidation-
reduction reactions.
 BIOLOGICAL OXIDATION-REDUCTION REACTIONS

 The free energy can be conserved and transformed as electrons are

transferred in oxidation-reduction reactions
 The flow of electrons can be coupled to do biological work

 Oxidation-reductions can be described as half-reactions:

 BIOLOGICAL OXIDATION-REDUCTION REACTIONS
Some terms:
 Reductant (Reducing agent): donate election

 Oxidant (Oxidizing agent): accept electron

 Redox pair (conjugate reductant-oxidant pair): electron donor and

acceptor in a half-reaction

Fe2+ and Fe3+ constitute a conjugate redox pair

 BIOLOGICAL OXIDATION-REDUCTION REACTIONS

Reduction potentials measure affinity of the electron acceptor of

each redox pair for electrons, referenced to the half reaction:

Standard reduction potential, E°

 Reactions with higher E° has greater ability to gain electrons

 REDUCTION POTENTIAL OF A REDOX PAIR
 Nernst equation:
E = Eº + RT/nF ln ([e- acceptor]/[e- donor])

 At room temperature (298 K or 25 oC):

E = Eº + (0.026V/n) ln ([e- acceptor]/[e- donor])

Where n is the number of electrons transferred

F is the Faraday constant (96480 J·V-1mol-1, or 23509 cal·V-1mol-1)
STANDARD REDUCTION POTENTIALS CAN BE USED
TO CALCULATE THE FREE-ENERGY CHANGE

Relationship between free energy and reduction potential

ΔG = -nF ΔE or ΔG’º = -nF ΔE’º
 provides a convenient way to measure ΔG of any biological oxidation
reactions
 BIOLOGICAL OXIDATIONS OFTEN INVOLVE
DEHYDROGENATION
The oxidation state of carbon in the biosphere
 In some cases, oxidation (loss of
electrons) is coincident with the
loss of hydrogen.
 Many biological oxidation
reactions are dehydrogenations
in which 1 or 2 hydrogen atoms
are transferred from a substrate
to a hydrogen acceptor.
 Many enzymes that catalyze
oxidation reactions are
dehydrogenases.
 CELLULAR OXIDATION OF GLUCOSE TO CO2
REQUIRES SPECIALIZED ELECTRON CARRIERS

 Series of controlled reactions, some of which are oxidations.

 Electrons removed in these oxidation steps are transferred to coenzymes
specialized for carrying electrons, such as NAD + and FAD
 ELECTRON CARRIERS

 NAD+ and NADP+

 FAD and FMN

 NAD+ AND NADP+
NADH and NADPH act with dehydrogenases as electron carriers.
Both of them are soluble, can be used by many different dehydrogenases.
 NAD+ AND NADP+
When NAD+ or NADP+ is reduced, the hydride ion could in principle be transferred
to either side of the nicotinamide ring: the front (A side) or the back (B side)
 NAD+ AND NADP+

 The (+) sign does not indicate the net charge of the molecules.
 Cellular concentration of NAD+ & NADH is ~10 μM.
 Cellular concentration of NADP+ & NDAPH is ~ 1 μM.
 The ratio of NAD+/NADH is high  used mostly in catabolic oxidation
 The ratio of NADP+/NADPH is low  used mostly in anabolic reduction
 The hydride ion (·H) on NADH or NADPH can be incorporated at either A or B
form. There’s no enzyme out there that can promiscuously catalyze both.
 NAD+ AND NADP+
The structure motif that binds NAD + or NADP+ in dehydrogenases is called the
“Rossmann fold”.

The nucleotide binding domain of the enzyme

lactate dehydrogenase.
NAD+ AND NADP+
 FAD AND FMN
 FAD and FMN are electron carriers that are tightly bound in flavoproteins.

Structures of oxidized and

reduced FAD and FMN
 FAD AND FMN
 Flavoproteins are enzymes that catalyze oxidation-reduction reactions using
either FMN or FAD as coenzyme

 Flavoproteins also serve as light receptors in cryptochromes and photolyases.

 OTHER ELECTRON CARRIERS

 Quinones
 Iron-sulfur proteins
 Cytochromes
 …
 SUMMARY
1. Bioenergetics and thermodynamics
Laws of thermodynamics
Quantitative relationships among free energy, enthalpy, and entropy
2. Phosphoryl group transfers and ATP
Roles of ATP in biological energy exchange
3. Biological oxidation-reduction reactions
Energetics of electron-transfer reactions
Electron carriers: NAD and NADP, FAD and FMN