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Bioenergetics
Quantitative study of energy transduction occurring in living cells.
Study of the nature and function of the chemical processes that are responsible for these energy transductions.
First Law of Thermodynamics
“For any physical or chemical change, the total amount of energy in the universe remains constant”
Therefore energy cannot be created or destroyed, but it can be converted from one form to another.
What is Gibbs Free energy?
Gibbs Free energy (G) is the amount of energy in a system available to do work.
Free energy changes derive from
Changes in heat content (DH) = enthalpy change
Changes in the state of order (DS) = entropy change
Negative Gibbs free energy
If ΔG is -ve, energy is liberated and the reaction is said to be exergonic
A reaction can occur spontaneously only if ΔG is –ve.
Positive Gibbs free energy
If ΔG = +ve
the reaction is said to be endergonic
For the reaction to occur it will require an energy input
Zero gibbs free energy
A system is at equilibrium and no net change can take place if ΔG is zero.
Is ΔG (Gibbs Free Energy) Dependent on the Path of the Reaction?
No, ΔG is independent of the path of the transformation.
Does ΔG Provide Information on the Rate of Reaction?
No, ΔG provides no information on the rate of the reaction. It only indicates whether a reaction is spontaneous or non-spontaneous. The rate of reaction depends on factors like activation energy, temperature, and catalysts, which ΔG does not address.
Why do we need energy?
Living organisms require continual input of free energy as many biological processes are endergonic:
Mechanical work e.g. Muscle contraction
Active transport
Synthesis of complex biomolecules from simple precursors
Also Signal transduction (environmental responses), generation of light (fire flies) and electricity (eels)
Role of ATP in Muscle Contraction
ATP provides energy for myosin head detachment from actin.
It re-cocks the myosin head for the next cycle.
ATP powers the active transport of calcium back into the sarcoplasmic reticulum for relaxation.
Without ATP, muscles stay contracted (rigor mortis).
Role of ATP in Active Transport
ATP provides energy to pump molecules against their concentration gradient through membrane transport proteins (e.g., Na+/K+ pump).
Na+/K+ pump: ATP hydrolysis moves 3 Na⁺ out and 2 K⁺ in, maintaining cell volume and resting potential.
Role of ATP in Neurotransmission
ATP powers the sodium-potassium pump to maintain resting membrane potential in neurons.
During neurotransmitter release, ATP is required for vesicle fusion with the presynaptic membrane and the release of neurotransmitters into the synapse.
Phototrophs
obtain energy by trapping light via photosynthesis
Chemotrophs
obtain energy by oxidation of food stuffs
Catabolism
Chemoorganotrophs
Extract energy from organic compounds by oxidation
Why should we have a controlled extraction of energy from food?
Regulation and control
Don’t want to release all the energy at once
Don’t want to increase body temperature excessively
Coupled reactions are more efficient
Not very mobile around body
Small carrier molecule better
Extraction of energy from food
Stage 1 - Large molecules broken down into smaller units. No useful energy captured
Stage 2 - Small molecules degraded into a few simple units that play a role in central metabolism. Some ATP generated.
Stage 3 - ATP produced from the complete oxidation of simple units by the final common pathways for oxidation of fuel.
What are fats broken down into?
Fatty acids and glycerol
What are polysaccharides broken down into?
glucose and other sugar
What are proteins broken down into?
amino acids
What Happens During the Degradation (Oxidation) of an Organic Compound?
As an organic compound is oxidized, electrons flow through intermediates.
The electrons are eventually transferred to oxygen (final electron acceptor) or are used to reduce other cellular components.
What Happens in a Redox Reaction?
In a redox reaction:
The electron donor is the reducing agent and is oxidized (loses electrons).
The electron acceptor is the oxidizing agent and is reduced (gains electrons).
Can Redox Reactions Do Work?
Yes, redox reactions involve electron flow, which can be harnessed to do work, similar to how electric circuits use electron flow to power devices.
Biological redox reactions
1. Direct electron transfer e.g.
Fe2+ + Cu2+ ↔ Fe3+ + Cu+
2. Direct transfer of hydrogen ions e.g.
AH2 + B ↔ A + BH2
3. Direct combination with oxygen as with mono-oxygenase reactions e.g.
R-CH3 + ½O2 ↔ R-CH2OH
4. The most common involve dehydrogenation
What is the Role of Dehydrogenases in Oxidation?
Dehydrogenases oxidize organic compounds by abstracting 2H⁺ and 2e⁻.
The electrons and protons are then passed to a mobile electron carrier.
This process occurs in biodegradation and energy abstraction (e.g., respiration).
What is the Role of Dehydrogenases in Reduction?
Dehydrogenases can reduce organic compounds by adding 2H⁺ and 2e⁻ from a mobile electron carrier.
This typically occurs in biosynthetic pathways (e.g., building complex molecules).
What is NADH and Its Role?
NADH is produced in catabolic reactions and by the TCA cycle.
It is used in the generation of ATP through oxidative phosphorylation (OxPhos).
NADH is usually found inside the mitochondria.
What is NADPH and Its Role?
NADPH is produced by the pentose phosphate pathway (PPP).
It is used primarily for reductive biosynthesis, such as fatty acid (FA) synthesis.
NADPH is usually found in the cytoplasm.
What is FADH₂ and Its Role?
FADH₂ is produced in catabolic reactions and by the TCA cycle.
It is used in the generation of ATP through oxidative phosphorylation (OxPhos), but generates less energy than NADH.
FADH₂ is usually found inside the mitochondria.
Why is ATP the energy currency of the cell?
Every living organism must generate ATP to survive.
When an organism stops producing ATP, it dies because ATP is essential for various cellular processes like metabolism, transport, and biosynthesis.
What Makes ATP an Energy-Rich Molecule?
ATP is an energy-rich molecule with high phosphoryl transfer potential.
It contains 2 phosphoanhydride bonds in its triphosphate unit, which store and release energy when broken.
ATP Hydrolysis Reaction 1:
ATP + H₂O → ADP + Pi + energy
The breakdown of ATP releases energy that can be used by the cell.
ATP Hydrolysis Reaction 2:
ATP + H₂O → AMP + PPi + energy
This reaction also releases energy and results in the production of AMP and pyrophosphate (PPi).
Is ATP Hydrolysis Thermodynamically Favorable?
Yes, free energy (ΔG) is negative, indicating ATP hydrolysis is thermodynamically unstable and spontaneous.
However, it is kinetically stable without a catalyst—its breakdown is slow unless catalyzed.
Free Energy of ATP Hydrolysis
he ΔG for ATP hydrolysis is:
-7.3 kcal/mol (for the conversion of ATP to ADP and Pi).
-10.9 kcal/mol (for the conversion of ATP to AMP and PPi).
Glucose Phosphorylation
Glucose + Pi → G-6-P
ΔG = +3.3 kcal/mol
This reaction is endergonic (requires energy input).
ATP Hydrolysis
ATP + H₂O → ADP + Pi
ΔG = -7.3 kcal/mol
This reaction is exergonic (releases energy).
Net Reaction (Glucose Phosphorylation Using ATP)
Glucose + ATP → G-6-P + ADP
ΔG = -4 kcal/mol
The overall reaction is exergonic, driven by ATP hydrolysis.
What is the Principal Mode of Energy Exchange in Biological Systems?
ATP is the principal immediate donor of free energy in biological systems.
ATP provides energy for cellular processes rather than serving as a long-term storage form.
What Happens to ATP After Its Formation?
ATP is consumed within minutes of formation to provide energy for various cellular activities.
ATP has a very high turnover—it is constantly being used and regenerated.
Diagram of ATP-ADP cycle
Substrate level phosphorylation
Transfer of phosphoryl group from metabolites with high-phosphoryl transfer potential to ADP producing ATP
Oxidative phosphorylation
Process of ATP formation as a result of transfer of electrons from fuels via electron carriers (NADH or FADH2) to the final electron acceptor oxygen.
In animals over 90% of ATP formed by this method. Carried out in the mitochondria.
What Are the Main Functions of Metabolism?
Metabolism is a highly coordinated cellular activity that serves four main functions:
Obtain energy (e.g., ATP) to fuel cellular processes.
Convert nutrients into the organism’s characteristic molecules.
Polymerize monomeric precursors (e.g., forming polysaccharides).
Synthesize and degrade molecules required for special cellular functions (e.g., intracellular messengers).
Characteristics of Central Metabolic Pathways
Hundreds of enzyme-catalyzed reactions occur in metabolism.
Central metabolic pathways are few in number but highly conserved throughout nature.
Two broad classes of metabolic pathways
CATABOLIC reactions-transform fuels into usable cellular energy
Degradative
Produces ATP
-ve free energy change
Produces reducing potential
Generates NADH + FADH2
ANABOLIC reactions-utilise the useful energy formed by catabolism to generate complex structures from simple ones.
Synthetic
Requires ATP
+ve free energy change
Requires reducing potential
Uses NADPH
Metabolism
Highly integrated network of chemical reactions, with thousands of reactions taking place.
The chemical reactions are linked to form interdependent metabolic pathways.
Pathways are regulated in common ways
Why is metabolic regulation required?
The human body requires energy for essential functions (e.g., breathing, circulating blood, walking).
The body does not have a constant external supply of energy.
Energy intake (food) is intermittent, usually 3–4 times a day.
Energy expenditure is continuous, with resting metabolism occurring all the time.
Occasional bursts of energy expenditure happen during physical activity.
The body must store energy and release it when required.
How does the body respond to extreme situations like exercise or illness that require sudden energy consumption?
The body increases its metabolic rate (up to 20 times the resting level) during situations like exercise or illness.
Despite reduced food intake, the body regulates the release of stored energy.
This helps meet the increased energy demand in these extreme situations.
What does metabolic regulation cover, and how does it work at a molecular level?
Covers the distribution and storage of nutrients after meals.
Manages the release of nutrients from stores.
Ensures nutrient delivery to tissues and utilization in individual cells as required.
Works at a molecular level, primarily by modulating enzyme activities.
How is metabolism regulated in the body?
Metabolism is regulated in three principal ways:
Levels and accessibility of substrates (through thermodynamics and compartmentation).
Amounts of metabolic enzymes (regulated by the rate of transcription and degradation).
Modulation of catalytic activities of enzymes (through allosteric regulation, covalent modification, and association with regulatory proteins).
What factors determine enzyme turnover in the body?
Enzyme turnover is determined by:
Alteration of transcription factor production in response to external signals.
Stability of mRNA species.
Rate of translation, which depends on various factors.
Rate of protein degradation.
Changes in the amount of enzyme present in the cell occur relatively slowly, ranging from minutes to hours.
How is enzyme activity modulated in metabolic pathways?
Metabolic pathways are interdependent.
Key enzymes, especially those at the rate-limiting or commitment steps, control the flow of substrates through a pathway.
These key enzymes can be regulated in several ways.
What is allosteric regulation and how does it work?
An allosteric enzyme has a site distinct from the substrate-binding site.
Ligands that bind to this site are called allosteric effectors or modulators.
Binding of these effectors causes conformational changes in the enzyme, altering its affinity for the substrate or other ligands.
Effectors can be positive (activators) or negative (inhibitors).
How does end-product or feedback inhibition work?
The end-product binds non-covalently to a specific regulatory site (allosteric site) of the enzyme.
Binding is dependent on the concentration of the end-product and the binding affinity.
This binding induces a conformational change in the enzyme, affecting the active site.
How do regulatory enzymes work?
Regulatory enzymes have several regulatory sites.
Each site selectively binds a ligand (either an activator or inhibitor).
The conformation of the enzyme’s active site reflects the summation of these signals.
Why are ATP levels always higher than ADP and AMP in the cell?
ATP levels are maintained higher than ADP and AMP to ensure efficient energy transfer and to drive cellular processes. The high ATP/ADP ratio promotes energy storage, while the AMP/ATP ratio signals when energy is low, triggering regulatory responses to conserve or generate energy.
How is metabolism controlled by adenylate control and energy charge?
Many reactions and pathways in metabolism are controlled by the energy status of the cell.
Energy charge ranges from 0 (all AMP) to 1 (all ATP).
ATP-generating pathways are inhibited by a high energy charge.
ATP-utilizing pathways are stimulated by a high energy charge.
The control of pathways has evolved to maintain energy charge within narrow limits, effectively buffering energy levels.
ATP-generating pathways catabolic
glycogenolysis
glycolysis
b-oxidation
ATP-utilising pathways anabolic
glycogenesis
gluconeogenesis
lipogenesis
purine + pyrimidine syntheses
What is covalent modification, and how does it affect enzyme regulation?
Covalent modification modifies the existing protein structure, allowing quicker regulation than changing enzyme levels (over seconds to minutes).
There are various types of covalent modification, including:
Adenylation
Methylation
Phosphorylation (the most common form)
How does the attachment of a functional group (like phosphate) to an amino acid side chain affect enzyme activity?
The attachment of a functional group (e.g., phosphate) is covalent and occurs selectively through an enzyme-catalyzed process.
This attachment induces a conformational change in the enzyme, which can alter its activity.
How does phosphorylation/dephosphorylation affect enzyme activity?
Phosphorylation/dephosphorylation alters the conformation of a protein, leading to:
Changes in Vmax and/or Km of the enzyme.
Altered sensitivity to substrates.
Altered sensitivity to inhibitors or activators.
The protein can be “locked” in a new conformation.
This process is reversible, which is necessary for regulation.
It is generally triggered by an external signal, leading to signal amplification.
What is the role of a phosphatase in enzyme regulation?
A phosphatase is an enzyme that removes phosphate groups from proteins (or other molecules) during dephosphorylation, reversing the effect of kinases and restoring the protein to its original state, which can alter its activity.
What is glycolysis?
A metabolic pathway that converts glucose to pyruvate. It is an ancient pathway used by a wide range of organisms from bacteria to humans.
Does glycolysis require oxygen?
No, glycolysis is anaerobic, meaning it does not require oxygen.
Where does glycolysis occur in eukaryotic cells?
In the cytosol.
Why is glucose important in cells?
Glucose is a key fuel in most cells, and in mammals, it is the only fuel that red blood cells use.
What are the two stages of glycolysis?
Stage 1: Trapping and destabilizing glucose to produce 2 x 3C molecules (requires 2 ATP per glucose).
Stage 2: Oxidation of the 3C molecules to pyruvate (produces 4 ATP and 2 NADH).
What is the first step in Stage 1 of glycolysis (Trapping and Destabilizing)?
Glucose enters cells via facillitated diffusion through specific transport proteins.
Once in the cell Glucose is trapped by phosphorylation.
Glucose 6-phosphate is negatively charge and cannot freely diffuse out of the cell.
Addition of the phosphate group begins the destabilisation process of glucose, which leads to further metabolism.
What type of enzyme is hexokinase, and what does it do?
Hexokinase is a kinase enzyme that can phosphorylate various hexose (six-carbon) sugars, including glucose, mannose, and even fructose.
What is the mechanism of action of hexokinase?
Hexokinase works through induced fit enzyme action, where the enzyme undergoes a conformational change upon substrate binding.
What is the equilibrium favoring in the hexokinase-catalyzed reaction?
The equilibrium of the reaction strongly favors the formation of glucose 6-phosphate, making the reaction effectively irreversible.
What type of regulation is hexokinase subject to?
Hexokinase is a regulatory enzyme in glycolysis and is inhibited by glucose 6-phosphate through feedback inhibition.
What is the second step in Stage 1 of glycolysis (Trapping and Destabilizing)?
Formation of Fructose 6-phosphate
Isomerisation of Glucose 6-P to Fructose 6-P is a completely reversible reaction carried out by the enzyme phosphoglucose isomerase.
Convert from one isomer (glucose) to another (fructose) by Tautomerisation
What is the third step in Stage 1 of glycolysis (Trapping and Destabilizing)?
Second phosphorylation reaction.
The enzyme Phosphofructokinase carries out this reaction.
Allosteric enzyme (Tetramer) which sets the pace of glycolysis
Inhibited by ATP, Citrate and H+ ions
Stimulated by AMP, ADP and Fruc 2,6-bisP
What are the fourth and fifth step in Stage 1 of glycolysis (Trapping and Destabilizing)?
Splitting Fructose 1,6-bisP into useful 3C fragments.
Cleavage of Fructose 1,6-bisP is catalysed by the enzyme Aldolase to yield 2 Triose phosphates
Readily reversible under normal physiological conditions
Is glyceraldehyde 3-phosphate (G3P) on the direct pathway of glycolysis?
Yes, G3P is on the direct pathway of glycolysis, while dihydroxyacetone phosphate (DHAP) is not.
Why does DHAP need to be converted into G3P in glycolysis?
DHAP needs to be converted into G3P to prevent the loss of a 3C fragment capable of generating ATP.
Which enzyme catalyzes the conversion of DHAP to G3P?
The enzyme triose phosphate isomerase (TIM) catalyzes the reversible conversion of DHAP to G3P.
What is the equilibrium between DHAP and G3P in glycolysis?
At equilibrium, 96% of the product is in the DHAP form, but the reaction is pushed towards G3P due to the subsequent steps in glycolysis and the removal of G3P.
Triose Phosphate Isomerase (TIM)
Great catalytic prowess, accelerates isomerisation by a factor of 1010 compared to simple base catalysis
Kinetically perfect enzyme, the rate limiting step is the diffusion-controlled encounter of substrate and enzyme.
So 2 molecules of G-3-P almost simultaneously from F 1,6-bisP
Stage 1 of Glycolysis summary
Glucose enters the cell via specific transporters.
Phosphorylation of Glucose traps it within the cell and begins the process of destabilisation
The 6C molecule is isomerised from an aldose to a ketose sugar prior to further destabilisation by phosphorylation.
The destabilised 6C sugar then fragments into two interconvertable 3C sugars.
STAGE 1 has utilised 2ATP molecules.
What is the sixth step of stage 2 of glycolysis (energy generation)?
Formation of a High Energy Bond
G 3-P is oxidised and phosphorylated by the enzyme G 3-P dehydrogenase.
Dehydrogenase transfer “high energy” electrons from complex organic molecule to NAD+ to form NADH
What are the two processes involved in the action of Glyceraldehyde 3-P dehydrogenase (GAPDH)?
Oxidation of the aldehyde group of G3P to a carboxylic acid by NAD+.
Joining of orthophosphate to the carboxylic acid, forming 1,3-bisphosphoglycerate, which is an acyl phosphate with high phosphoryl-transfer potential.
What is the seventh step of stage 2 of glycolysis (energy generation)?
ATP generation from 1,3-bisPglycerate
Substrate level Phosphorylation
Remember Glucose (6C) yields 2 x 3C intermediates therefore 2 ATP’s generated per glucose molecule.
What are the eighth, ninth and tenth steps of stage 2 of glycolysis (energy generation)?
Generation of additional ATP and pyruvate formation (2 per glucose molecule)
Phosphoryl group on 3-Pglycerate shifts position, followed by dehydration and formation of a C=C bond.
Increases transfer potential of phosphoryl group.
What is the role and regulation of Pyruvate Kinase in glycolysis?
Pyruvate kinase catalyzes the irreversible transfer of a phosphoryl group to form ATP through substrate-level phosphorylation.
It is a regulatory enzyme activated by Fructose 1,6-bisphosphate and inhibited by ATP and alanine.
Stage 2 of glycolysis summary
What is the overall reaction of glycolysis?
Glucose + 2 NAD+ + 2 ADP + Pi → 2 Pyruvate + 2 NADH + 2 ATP + 2 H+ + 2 H2O
Why must NAD+ be replenished during glycolysis?
Glycolysis would not proceed for long if pyruvate were the final metabolite, as it would disturb the redox balance. NAD+ is limited in cells and must be replaced to maintain the process.
What happens to NAD+ during glycolysis and why is it important?
During glycolysis, NAD+ is converted to NADH. Glycolysis cannot continue if NAD+ levels decrease, as NAD+ is needed for the oxidation of G3P in step 6.
What happens under aerobic conditions to regenerate NAD+?
Under aerobic conditions, electrons from NADH are transferred to oxygen via the electron transport chain to form H2O, ATP, and regenerate NAD+.
What happens under anaerobic conditions to regenerate NAD+?
Under anaerobic conditions, electrons from NADH are transferred to pyruvate, forming lactate or ethanol, and regenerating NAD+ for glycolysis to continue.
What is ethanol formation, and where does it occur?
Ethanol formation occurs in yeast and some microorganisms. It is an anaerobic process where NAD+ is regenerated to allow glycolysis to continue.
What is lactic acid formation, and when does it occur?
Lactic acid formation occurs in microorganisms and higher organisms when oxygen is limited, such as during intense exercise, to regenerate NAD+ for glycolysis.
What is the overall reaction for ethanol formation during anaerobic glycolysis?
Glucose + 2 H+ + 2 ADP + 2 Pi → 2 Ethanol + 2 CO2 + 2 ATP + 2 H2O
How is NAD+ regenerated under aerobic conditions when NADH cannot enter the mitochondria?
NADH generated during glycolysis cannot enter the mitochondria directly, so NAD+ is regenerated indirectly through oxidative phosphorylation (OX PHOS) using specific shuttles.
What is the fate of pyruvate under aerobic conditions?
Under aerobic conditions, pyruvate enters the mitochondria, where it is oxidized to acetyl-CoA, which then enters the TCA cycle for further energy extraction.
Malate/Aspartate Shuttle
