13 | Bioenergetics and Reactions

© 2013 W. H. Freeman and Company

 CHAPTER 13
Bioenergetics and Reactions

Key topics:
– Thermodynamics applies to biochemistry, too
– Organic chemistry principles are still valid
– Some biomolecules are “high energy” with respect
to their hydrolysis and group transfers
– Energy stored in reduced organic compounds can be
used to reduce cofactors such as NAD+ and FAD,
which serve as universal electron carriers
 Life needs energy

• Recall that living organisms are built of complex

structures
• Building complex structures that are low in entropy is
only possible when energy is spent in the process
• The ultimate source of this energy on Earth is the
sunlight
Metabolism is the sum of all chemical
reactions in the cell

• Series of related reactions form metabolic pathways

• Some pathways are primarily energy-producing
– This is catabolism
• Some pathways are primarily using energy to build
complex structures
– This is anabolism or biosynthesis
 Laws of thermodynamics
apply to living organisms
• Living organisms cannot create energy from nothing
• Living organisms cannot destroy energy into nothing
• Living organism may transform energy from one form to another

• In the process of transforming energy, living organisms

must increase the entropy of the universe
• In order to maintain organization within themselves,
living systems must be able to extract useable energy
from their surroundings, and release useless energy
(heat) back to their surroundings
• 1st law : For any physical or chemical change,
the total amount of energy in the universe
remains constant

• 2nd law : In all natural processes, the entropy

of the universe increases
Gibbs free energy

G: Gibbs free energy

H: Enthalpy
S: Entropy
Free energy of reactions
To derive the Gibbs free energy equation for an isolated system, let Stot be the total entropy of the isolated system, that is, a system which
cannot exchange heat or mass with its surroundings. According to the second law of thermodynamics:

          
Now consider diabatic systems, having internal entropy Sint. Such a system is thermally connected to its surroundings, which have
entropy Sext. The entropy form of the second law does not apply directly to the diabatic system, it only applies to the closed system
formed by both the system and its surroundings. Therefore a process is possible if

                                        .
We will try to express the left side of this equation entirely in terms of state functions. ΔSext is defined as:

                              
Temperature T is the same for two systems in thermal equilibrium. ΔQ is heat transferred to the system, so − ΔQ is heat transferred to
the surroundings, and −ΔQ/T is entropy gained by the surroundings. We now have:

                                   

Multiply both sides by T:

                                      
ΔQ is heat transferred to the system; if the process is now assumed to be isobaric, then ΔQp = ΔH:
                                        
ΔH is the enthalpy change of reaction (for a chemical reaction at constant pressure and temperature). Then
                                        
for a possible process. Let the change ΔG in Gibbs free energy be defined as

Notice that it is not defined in terms of any external state functions, such as ΔSext or ΔStot. Then the second law becomes, which also tells
us about the spontaneity of the reaction:
                  favored reaction (Spontaneous)
                  Neither the forward nor the reverse reaction prevails (Equilibrium)
                  disfavored reaction (Nonspontaneous)
Gibbs free energy and equilibrium constant
Free energy, or the equilibrium constant,
measure the direction of processes
Energetics of Some Chemical Reactions
• Hydrolysis reactions tend to be strongly favorable
(spontaneous)
• Isomerization reactions have smaller free-energy
changes
– Isomerization between enantiomers: G = 0
• Complete oxidation of reduced compounds is
strongly favorable
– This is how chemotrophs obtain most of their energy
– In biochemistry the oxidation of reduced fuels with O2 is
stepwise and controlled
– Recall that being thermodynamically favorable is not the
same as being kinetically rapid
Energetics within the cell are not standard
• The actual free-energy change of a reaction in the
cell depends on:
– The standard change in free energy
– Actual concentrations of products and reactants
– For the reaction aA + bB cC + dD:
[C ]c [ D ]d
G  G '  RT ln
[ A]a [ B ]b
• Standard free-energy changes are additive:
(1) A  B ΔG’1
(2) B  C ΔG’2
Sum: A  C ΔG’1 + ΔG’2
 Lesson in Quantum Chemistry
• Most organic molecules, including the reduced fuels, are in
the singlet spin state
– All electrons are paired into electron pairs
• Molecular oxygen is in the triplet spin state
– Two electrons are unpaired
• Direct electron transfer from a singlet reduced species to a
triplet oxidizing species is quantum-mechanically forbidden
• This is why direct oxidation (spontaneous combustion) of
biomolecules does not occur readily
• Few cofactors, such as transition metal ions, and flavin
adenine dinucleotide are able to catalyze consecutive single-
electron transfers needed for utilization of O2
 Review of Organic Chemistry

• Most reactions in biochemistry are thermal

heterolytic processes
• Nucleophiles react with electrophiles
• Heterolytic bond breakage often gives rise to
transferable groups, such as protons
• Oxidation of reduced fuels often occurs via transfer of
electrons and protons to a dedicated redox cofactor
 Chemical Reactivity
Most reactions fall within few categories:
•Cleavage and formation of C–C bonds
•Cleavage and formation of polar bonds
‒ Nucleophilic substitution mechanism
‒ Addition–elimination mechanism
• Hydrolysis and condensation reactions
•Internal rearrangements
•Eliminations (without cleavage)
•Group transfers (H+, CH3+, PO32–)
•Oxidations-reductions (e– transfers)
 Chemistry at Carbon

• Covalent bonds can be broken in two ways

• Homolytic cleavage is very rare
• Heterolytic cleavage is common, but the products
are highly unstable and this dictates the chemistry
that occurs
Homolytic vs. Heterolytic Cleavage
Nucleophiles and Electrophiles
in Biochemistry
Examples of Nucleophilic Carbon-Carbon
Bond Formation Reactions
Isomerizations and Eliminations:
No Change in Oxidation State
 Addition–Elimination Reactions

• Substitution from sp3 carbon proceeds normally via

the nucleophilic substitution (SN1 or SN2) mechanism
• Substitution from the sp2 carbon proceeds normally
via the nucleophilic addition–elimination mechanism
– Nucleophile adds to the sp2 center giving a
tetrahedral intermediate
– Leaving group eliminates from the tetrahedral
intermediate
– Leaving group may pick up a proton
Addition–Elimination Reactions
 Group Transfer Reactions

• Proton transfer, very common

• Methyl transfer, various biosyntheses
• Acyl transfer, biosynthesis of fatty acids
• Glycosyl transfer, attachment of sugars
• Phosphoryl transfer, to activate metabolites
‒ also important in signal transduction
 Nucleophilic Displacement

• Substitution from sp3 phosphorous proceeds via the

nucleophilic substitution (usually associative, SN2-like)
mechanism
– Nucleophile forms a partial bond to the
phosphorous center giving a pentacovalent
intermediate or a pentacoordinated transition state
Nucleophilic Displacement
 Phosphoryl Transfer from ATP
ATP is frequently the donor of the phosphate in the
biosynthesis of phosphate esters.
Hydrolysis of ATP is highly favorable
under standard conditions
• Better charge separation
in products
• Better solvation of
products
• More favorable resonance
stabilization of products
 Actual G of ATP hydrolysis
differs from G’
• The actual free-energy change in a process depends on:
– The standard free energy
– The actual concentrations of reactants and products
• The free-energy change is more favorable if the reactant’s
concentration exceeds its equilibrium concentration
• True reactant and the product are Mg-ATP and Mg-ADP,
respectively
[MgADP ]  [Pi ]
 G also Mg++ dependent G  G ' RT ln
[MgATP 2 ]
G of ATP hydrolysis is
Mg++ dependent
Cellular ATP concentration is usually far above the equilibrium
concentration, making ATP a very potent source of chemical energy.
Several phosphorylated compounds have large
G’ for hydrolysis
• Again, electrostatic repulsion within the reactant
molecule is relieved
• The products are stabilized via resonance, or by more
favorable solvation
• The product undergoes further tautomerization
Phosphates: Ranking by the Standard Free
Energy of Hydrolysis
Phosphate can be transferred from compounds with
higher ΔG’ to those with lower ΔG’.

Reactions such as

PEP + ADP =>

Pyruvate + ATP

are favorable, and

can be used to
synthesize ATP.
 Hydrolysis of Thioesters

• Hydrolysis of thioesters is strongly favorable

– such as acetyl-CoA
• Acetyl-CoA is an important donor of acyl groups
– Feeding two-carbon units into metabolic pathways
– Synthesis of fatty acids
• In acyl transfers, molecules other than water accept
the acyl group
Hydrolysis of Thioesters
Molecular Basis for Thioester Reactivity
The orbital overlap between the carbonyl group and
sulfur is not as good as the resonance overlap between
oxygen and the carbonyl group in esters.
 Oxidation-Reduction Reactions
Reduced organic compounds serve as fuels from which
electrons can be stripped off during oxidation.
 Reversible Oxidation of
a Secondary Alcohol to a Ketone
• Many biochemical oxidation-reduction reactions
involve transfer of two electrons
• In order to keep charges in balance, proton transfer
often accompanies electron transfer
• In many dehydrogenases, the reaction proceeds by a
stepwise transfers of proton (H+) and hydride (:H–)
 Reduction Potential
• Reduction potential (E)
– Affinity for electrons; higher E, higher affinity
– Electrons transferred from lower to higher E

E’ = -(RT/nF)ln (Keq) = G’/nF

∆E’ = E’(e- acceptor) – E’(e- donor)

∆G’ = –nF∆E’
For negative G need positive E
E(acceptor) > E(donor)
 NAD and NADP are
common redox cofactors
• These are commonly called pyridine nucleotides
• They can dissociate from the enzyme after the
reaction
• In a typical biological oxidation reaction, hydride
from an alcohol is transferred to NAD+ giving NADH
 NAD and NADP are
common redox cofactors
 Formation of NADH can be monitored
by UV-spectrophotometry
• Measure the change of absorbance at 340 nm
• Very useful signal when studying the kinetics of
NAD-dependent dehydrogenases
 Flavin cofactors allow
single electron transfers
• Permits the use of molecular oxygen as an ultimate electron acceptor
– flavin-dependent oxidases
• Flavin cofactors are tightly bound to proteins
 Chapter 13: Summary
In this chapter, we learned:
• The rules of thermodynamics and organic chemistry still apply to living
systems
• Reactions are favorable when the free energy of products is much
lower than the free energy of reactants
• Biochemical phosphoryl transfer reactions are favorable when:
– The phosphate donors are destabilized by electrostatic repulsion,
– and the reaction products are often stabilized by resonance
• Unfavorable reactions can be made possible by chemically coupling a
highly favorable reaction to the unfavorable reaction
• Oxidation-reduction reactions commonly involve transfer of electrons
from reduced organic compounds to specialized redox cofactors
– Reduced cofactors can be used in biosynthesis, or may serve as a source of energy for ATP
synthesis