DBT2118: Biochemistry (II)
Lecture 1
Introduction to metabolism & Common reactions
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This course is aimed for you to be able to understand the major biochemicals
processes that occur in biology
• Biological energy generation & storage
• Biochemical synthesis and degradation of biomolecules
(sugars, lipids, amino acids, nucleic acids)
Reference material
Biochemistry 4th edition, Mathews, Van Holde, Appling, Anthony‐Cahill. Pearson ISBN:978‐0‐13‐800464‐4
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Lehninger Principles of Biochemistry 4th edition, David L. Nelson, Michael M. Cox. W. H. Freeman ISBN:978‐0716743392
Metabolism
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Metabolism is the study of chemical reactions and their regulations inside a cell…
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Topic
Carbohydrate Metabolism
Carbohydrate Metabolism
Citric Acid Cycle and Glyoxylate Cycle
Citric Acid Cycle and Glyoxylate Cycle
Electron Transport, Oxidative Phosphorylation
Exam 1
Photosynthesis
Photosynthesis
Lipid Metabolism I: Fatty acid
Lipid Metabolism I: Fatty acid
Interorgan and Intracellular Coordination
Lipid Metabolism II: Membrane
Lipids/Exam2
Metabolism of Nitrogenous Compounds I
Metabolism of Nitrogenous Compounds I
Metabolism of Nitrogenous Compounds II
Metabolism of Nitrogenous Compounds II
Nucleotide Metabolism
Exam 3
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Topic
Carbohydrate Metabolism
Carbohydrate Metabolism
Citric Acid Cycle and Glyoxylate Cycle
Citric Acid Cycle and Glyoxylate Cycle
Electron Transport, Oxidative Phosphorylation
Exam 1
Photosynthesis
Photosynthesis
Lipid Metabolism I: Fatty acid
Lipid Metabolism I: Fatty acid
Interorgan and Intracellular Coordination
Lipid Metabolism II: Membrane
Lipids/Exam2
Metabolism of Nitrogenous Compounds I
Metabolism of Nitrogenous Compounds I
Metabolism of Nitrogenous Compounds II
Metabolism of Nitrogenous Compounds II
Nucleotide Metabolism
Exam 3
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Catabolism vs. Anabolism
• Catabolism: reactions that break down nutrients and collect released energy and reducing power
• Catabolic pathways are convergent
• Anabolism: reactions that synthesize needed
compounds, using stored energy and reducing
power
• Anabolic pathways are divergent
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Converging Catabolism vs. Diverging Anabolism
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Metabolic currency ($$$)
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Metabolic intermediates serves as “currency” to convert one type of molecule to another.
NADH NADPH FADH2
Electron carriers
Participates in Oxidation and reductions
ATP = Energy carrier
Used to increase thermodynamic favorability
of reaction 7
Today’s topics:
1. ATP & other high energy phosphates
2. Oxidation & reduction (NADH, NADPH, FADH2)
3. Common reactions in biochemistry
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ATP & High energy phosphate donors
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The adenylates (ATP, ADP, AMP) are the primary energy currency
High in energy!!
• The fundamental biological role of ATP as an energy‐coupling compound
is to convert thermodynamically unfavorable processes into favorable
processes.
• Activated intermediates, such as ATP, allow reactions to occur under
physiologically relevant concentrations of metabolic intermediates.
• Reduced charge repulsion in products
• Better resonance stabilization of products
• More favored solvation of products
 ΔG'° is ‐30.5 kJ/mol0
ATP + H2O  ADP + Pi is VERY favorable reaction
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Resonance makes Phosphate (Pi or PO4)very stable
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The ΔG of ATP hydrolysis is large and negative
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ATP is used to DRIVE thermodynamically unfavorable reactions
Pi
ATP
ADP
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ΔG'° values of phosphate hydrolysis reflect ‘phosphoryl transfer potential’ (ptp)
High ptp
Low ptp
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Other ‘high‐energy phosphate’ compounds have great stabilization of hydrolysis products
Reduced charge repulsion and tautomerization:
(Phosphoenolpyruvate)
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Other ‘high‐energy phosphate’ compounds have great stabilization of hydrolysis products
Reduced charge repulsion and resonance stabilization:
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Thioesters also serve as energy currencies
Resonance of carboxylic acid stabilizes the product
Some bacteria can use acetyl‐
CoA to produce acetate + ATP
9.8 kJ/mol
Pi
CoA‐SH
phosphate acetyltransferase
Acetyl‐Phosphate
Acetate kinase
ADP
ATP
‐13 kJ/mol
Similar ΔG'° of hydrolysis as ATP 16
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Common functional groups in biochemistry
Acetyl‐CoA is a common metabolite used to both synthesize lipids and respiration
*note: Metabolites = chemical compounds found in the cell
Acetyl‐
CoA
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Oxidation & Reductions
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Simple oxidation and reduction (Redox) in common context of metabolism:
Most reduced
Most oxidized
CH4
CO2
methanol
formaldehyde
Formic acid
(or formate)
oxidation
Reduction
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Redox energy currencies transfer reducing power (ex: NAD and NADP)
• 2 electron, 1 proton carriers
• cosubstrates: diffuse between
different enzymes
• NAD: primarily used in
catabolism
• NADP: primarily used in
anabolism
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NAD+ accepts a “hydride” to become NADH
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Cells use NADH as a reducing agent to reduce metabolites
Oxidation/reduction: NADH dependent Lactate dehydrogenase
Hydride transfer (H:‐) from NADH to Pyruvate, forming lactate
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NADH vs. NADPH
• NAD+ is the cofactor for most enzymes that act in the direction of substrate oxidation
(dehydrogenases).
• NADPH usually functions as a cofactor for reductases, enzymes that catalyze substrate
reduction.
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FAD and FMN are other redox currencies
FAD: Flavin adenosine dinucleotide
• Usually prosthetic groups: tightly bound to enzyme
• Can transfer 1 or 2 electrons (plus 1 or 2 protons)
FMN: Flavin mononucleotide
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FAD and FMN are other redox currencies
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Common functional groups & reactions
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Common functional groups in biochemistry
Functional groups are groups of atoms added to carbon skeleton that have specific properties.
Carbon functional groups
Oxygen functional groups
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Common functional groups in biochemistry
Sulfur functional groups
Nitrogen functional groups
Phosphor functional groups
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Carbonyl molecules
• Carbonyl chemistry account for a large percentage of biochemical
reactions because the vast majority of biological molecules contain them.
• Most of the chemistry of carbonyl groups involves nucleophiles
(abbreviated “Nu:”) and electrophiles.
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Common biochemical nucleophiles & electrophiles
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Oxyanion mechanism
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Oxyanion chemistries occur frequently to reactions involving carbonyls.
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A nucleophile attacks the carbonyl center (electrophile), generating a tetrahedral
oxyanion intermediate.
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In the case of a substitution reaction, when negative charge on oxygen comes back down, the –OR group leaves the molecule, completing the reaction.
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Oxyanion chemistries
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α‐carbon of carbonyl has slightly acidic proton (α‐hydrogen)
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α‐hydrogen is weakly acidic proton
(compared to other C‐H which is not acidic) that can be pulled off by a base.
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It is weakly acidic because its conjugate base is stabilized via enolate resonance.
Electron can delocalize
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Enolate intermediate enables Aldol and Claisen condensation •
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Forming Carbon – Carbon Bonds
Used in fatty acid metabolism
Carbonyl condensation reactions: • These reactions are initiated by
deprotonation of the weakly acidic α‐
hydrogen to give a resonance‐stabilized
enolate ion (top).
• Aldol condensation (left): the enolate
adds to an aldehyde or ketone, yielding
a b‐hydroxy carbonyl product.
• Claisen condensation (right side): the
enolate adds to an ester (can also be
CoA‐thioester), yielding a b‐keto
product.
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Elimination reaction
Eliminations:
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Dehydration of β‐hydroxyl group
• Dehydration of β‐hydroxyl group is a common
mechanism used in:
• fatty acid biosynthesis
• TCA cycle
• Amino acid biosynthesis
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Oxidation/Reduction reactions (Redox)
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Energy production in most cells involves the oxidation of fuel molecules such as
glucose and fats.
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Oxidation‐reduction, or redox, chemistry is the core of metabolism. •
Redox reactions involve reversible electron transfer from a donor (the reductant) to
an acceptor (the oxidant).
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In below example, because the alcohol has lost a pair of electrons and two hydrogen
atoms (essentially H2),
• this type of oxidation is called dehydrogenation, and enzymes that catalyze this
reaction are called dehydrogenase. NADH is called reducing cofactor, reducing
equivalent, or sometimes reducing power
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Pathway regulation typically at large ΔG steps
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Enzyme regulation and control
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To avoid such events from happening, biology evolved regulations at various levels
(transcriptional, translational, protein and substrate levels such as allosteric inhibitions)
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Regulations typically are placed on enzymatic steps with large ΔG
A reaction with small or zero ΔG:
A reaction with large ΔG:
Can’t control anything…
Control whether or not reaction occurs is meaningful
Both pathways are favorable, so cell can control when to turn on which pathway…
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Enzyme regulation and control
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While cells have transcriptional and translational controls over gene expression and protein
translation, it is also necessary to have protein level regulation to ensure proper balance of
cellular resources.
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Substrate level control:
• Product of a reaction may serve as an inhibitor to its enzyme
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Feedback controls:
• Metabolites downstream of a pathway can inhibit upstream enzymes of the same
metabolic pathway
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Feedback controls
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Most frequently, negative feedback controls are used to control flow of metabolites
inside a cell
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Negative feedback (product feedback inhibition): product of a pathway inhibits
its upstream
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Activation feedback: product of a pathway may activate some other pathways
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https://www.youtube.com/watch?v=DHZtOKyMPRY
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How are downstream products able to control upstream enzymes?
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Allosteric regulations – interactions between enzyme and a molecule that induces
enzyme to undergo conformational change.
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Conformational change may increase enzyme activity or decrease.
Example: ACTase
• ATP is an activator.
• CTP is an inhibitor.
*Cytidine is a pyrimidine
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Enzyme regulations: covalent modifications
Covalent modifications are ways to “irreversibly” inhibit an enzyme
This covalent modification can be removed by another enzyme
(for example, kinase for phosphorylation & phosphatase for de‐phosphorylation)
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Reversible covalent modification by kinases/phosphatases: •
The target residues for ATP‐dependent phosphorylation by kinases are serine, threonine, or
tyrosine.
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The phosphoprotein is dephosphorylated by a phosphatase‐catalyzed hydrolysis reaction.
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How we know enzyme’s participation in a pathway
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By inactivating individual enzymes, mutations and enzyme inhibitors
help identify the metabolic roles of
enzymes.
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The steps of a hypothetical
metabolic pathway are identified by
analysis of mutants defective in
individual steps of the pathway.
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We can identify metabolite C as the
substrate for enzyme III by the
absence of this enzyme in mutants
that accumulate C.
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We know that D and E follow C in
the pathway because feeding either D or E to mutants defective in
enzyme III bypasses the genetic
block and allows the cells to grow.
(*note: provided that cell needs metabolite E to grow)
*note
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Overview of Glycolysis
Payoff phase
Embden‐Meyerhof‐Parnas (EMP) pathway
Investment phase
hexokinase
Phosphoglucoisomerase
Phosphofructokinase
Glyceraldehyde‐3‐phosphate
dehydrogenase
Phosphoglycerate kinase
Phosphoglycerate mutase
enolase
aldolase
Pyruvate kinase
Triose isomerase
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