Download Principles of BIOCHEMISTRY - Valdosta State University

Introduction to Metabolism
• A hummingbird has a
rapid rate of
metabolism, but its
basic metabolic
reactions are the same
as those in many
diverse organisms
• Autotrophs – use CO2 as sole carbon
source (plants, photosynthetic
bacteria, etc.)
• Heterotrophs-obtain carbon from
their environment
• Constant cycling of material between
autotrophs and heterotrophs
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Metabolism Is the Sum of Cellular Reactions
• Metabolism - the entire network of chemical
reactions carried out by living cells
• Metabolites - small molecule intermediates in
the degradation and synthesis of polymers
• Catabolic reactions - degrade molecules to
create smaller molecules and energy
• Anabolic reactions - synthesize molecules for
cell maintenance, growth and reproduction
Anabolism and catabolism
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Metabolic Reactions
• Metabolism includes all enzyme catalyzed
reactions
• The metabolism of the four major groups of
biomolecules will be considered:
Carbohydrates
Lipids
Amino Acids
Nucleotides
Organization of Metabolic Reactions
• Occur via pathways – series of organized
reaction steps
• Compartmentalized – certain reactions occur
in particular cells, organelles or other specific
sites
• Pathways are regulated – controlled
– to keep anabolism and catabolism reactions
separate (some use the same enzymes)
– Timing to produce products only when necessary
– At least one step in a pathway needs to be
irreversible (exergonic, -G)
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Types of pathways
• Individual reaction series
– Linear (can branch out)
– Cyclic
– Spiral
• Connecting pathways
– Converging (metabolic)
– Diverging (anabolic)
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Forms of metabolic pathways
(a)Linear
or branched
(b) Cyclic
(c) Spiral pathway
(fatty acid
biosynthesis)
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Metabolism Proceeds by Discrete Steps
• Multiple-step pathways permit control of
energy input and output
• Catabolic multi-step pathways provide
energy in smaller stepwise amounts)
• Each enzyme in a multi-step pathway usually
catalyzes only one single step in the pathway
• Control points occur in multistep pathways
• Single-step vs multistep pathways
• A multistep enzyme
pathway releases
energy in smaller
amounts that can be
used by the cell
Metabolic Pathways Are Regulated
• Regulation permits response to changing
conditions
• Common ways to regulate
•
(1) Supply of substrates(concentration)
(2) Removal of products
(3) Pathway enzyme activities
•Allosteric regulation
•Covalent modification
Feedback inhibition
• Product of a pathway controls the rate of its own
synthesis by inhibiting an early step (usually the
first “committed” step (unique to the pathway)
Feed-forward activation
• Metabolite early in the pathway activates an
enzyme further down the pathway
Covalent modification for enzyme regulation
• Interconvertible enzyme activity can be rapidly and
reversibly altered by covalent modification
• Protein kinases phosphorylate enzymes (+ ATP)
• Protein phosphatases remove phosphoryl groups
• The initial signal may be amplified by the “cascade”
nature of this signaling
Reaction Types in Pathways
1. Oxidation-Reduction (Redox)
2. Making or breaking C-C bonds
3. Internal rearrangements,
isomerizations or eliminations
4. Group transfers
5. Free radical reactions
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Redox reactions
• Oxidation – loss of electrons, gain of
oxygen, loss of hydrogen
– Hydrogenases
– Oxidases
• Note the different oxidation states of
carbon
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Carbon-Carbon Bonds
• Bond cleavage
– Homolytic (1 electron for each atom)
– Heterolytic (both electrons to one atom)
– Recall
• Nucleophiles (attracted to + charges)
• Electrophiles (attracted to – charges)
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Common Reaction Types
• Many use the carbonyl group C=O
• + on Carbon; - on Oxygen
– Reactive group in
• Aldol condensations
• Claisen condensations
• Decarboxylations
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Internal Reactions
• Rearrangements, isomerizations,
eliminations
– Groups
– Bonds
– Atoms
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Group Transfers
• There are many groups to transfer
– Acyl
– Glycosyl
– Phosphoryl
• Phosphate = Pi
• Pyrophosphate = PPi
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Free Radicals
• Unpaired electrons
• More common than previously thought
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10.3 Major Pathways in Cells
• Metabolic fuels
Three major nutrients consumed by mammals:
(1) Carbohydrates - provide energy
(2) Proteins - provide amino acids for protein
synthesis and some energy
(3) Fats - triacylglycerols provide energy and
also lipids for membrane synthesis
Fig 10.5
• Overview of
catabolic
pathways
Catabolism produces compounds
for energy utilization
• Three types of compounds are produced that
mediate the release of energy
(1) Acetyl CoA
(2) Nucleoside triphosphates (e.g. ATP)
(3) Reduced coenzymes (NADH, FADH2, QH2)
Reducing Power
• Electrons of reduced coenzymes flow toward O2
• This produces a proton flow and a transmembrane
potential
• Oxidative phosphorylation is the process by
which the potential is coupled to the reaction:
ADP + Pi
ATP
10.4 Compartmentation and
Interorgan Metabolism
• Compartmentation of metabolic processes permits:
- separate pools of metabolites within a cell
- simultaneous operation of opposing metabolic
paths
- high local concentrations of metabolites
- coordinated regulation of enzymes
• Example: fatty acid synthesis enzymes (cytosol),
fatty acid breakdown enzymes (mitochondria)
Fig. 10.6 Compartmentation of
metabolic processes
10.5 Thermodynamics and Metabolism
A. Free-Energy Change
• Free-energy change (G) is a measure of the
chemical energy available from a reaction
G = Gproducts - Greactants
• H = change in enthalpy
• S = change in entropy
Relationship between energy and entropy
• Both entropy and enthalpy contribute to G
G = H - TS
(T = degrees Kelvin)
-G = a spontaneous reaction in the
direction written
+G = the reaction is not spontaneous
G = 0 the reaction is at equilibrium
The Standard State (Go) Conditions
• Reaction free-energy depends upon conditions
• Standard state (Go) - defined reference conditions
Standard Temperature = 298K (25oC)
Standard Pressure = 1 atmosphere
Standard Solute Concentration = 1.0M
• Biological standard state = Go’
Standard H+ concentration = 10-7 (pH = 7.0) rather
than 1.0M (pH = 1.0)
B. Equilibrium Constants and
Standard Free-Energy Change
• For the reaction: A + B
C+D
Greaction = Go’reaction + RT ln([C][D]/[A][B])
• At equilibrium: Keq = [C][D]/[A][B] and
Greaction = 0, so that:
Go’reaction = -RT ln Keq
C. Actual Free-Energy Change Determines
Spontaneity of Cellular Reactions
• When a reaction is not at equilibrium, the
actual free energy change (G) depends
upon the ratio of products to substrates
• Q = the mass action ratio
G = Go’ + RT ln Q
Where Q = [C]’[D]’ / [A]’[B]’
10.6 The Free Energy of ATP
• Energy from oxidation of metabolic fuels is
largely recovered in the form of ATP
Table 10.1
Fig 10.7
• Hydrolysis of
ATP
Fig 10.8 Complexes between ATP and Mg2+
ATP is an “energy-rich” compound
• A large amount of energy is released in the
hydrolysis of the phosphoanhydride bonds of
ATP (and UTP, GTP, CTP)
• All nucleoside phosphates have nearly equal
standard free energies of hydrolysis
Energy of phosphoanhydrides
(1) Electrostatic repulsion among negatively
charged oxygens of phosphoanhydrides of ATP
(2) Solvation of products (ADP and Pi) or (AMP
and PPi) is better than solvation of reactant ATP
(3) Products are more stable than reactants
There are more delocalized electrons on ADP, Pi
or AMP, PPi than on ATP
10.7 The Metabolic Roles of ATP
• Energy-rich compounds can drive biosynthetic
reactions
• Reactions can be linked by a common energized
intermediate (B-X) below
A-X + B
A + B-X
B-X + C
B + C-X
Glutamine synthesis requires ATP energy
A. Phosphoryl-Group Transfer
• Phosphoryl-group-transfer potential - the ability
of a compound to transfer its phosphoryl group
• Energy-rich or high-energy compounds have
group transfer potentials equal to or greater than
that of ATP
• Low-energy compounds have group transfer
potentials less than that of ATP
Table 10.3
B. Production of ATP by
Phosphoryl-Group Transfer
• Metabolites with high phosphoryl-group-transfer
potentials can donate a phosphoryl group to ADP
to form ATP
• Energy-rich compounds are intermediates in
catabolic pathways
• Energy storage compounds can be energy-rich
Fig 10.9 Relative phosphoryl-grouptransfer potentials
Fig 10.10 Transfer of the phosphoryl
group from PEP to ADP
• Phosphoenolpyruvate (PEP) (a glycolytic
intermediate) has a high P-group transfer potential
• PEP can donate a P to ADP to form ATP
Phosphagens: Energy-rich storage
molecules in animal muscle
• Phosphocreatine (PC) and phosphoarginine (PA)
are phosphoamides
• Have higher group-transfer potentials than ATP
• Produced in muscle during times of ample ATP
• Used to replenish ATP when needed via creatine
kinase reaction
Fig 10.11 Structures of PC and PA
C. Nucleotidyl-Group Transfer
• Transfer of the nucleotidyl group from ATP is
another common group-transfer reaction
• Synthesis of acetyl CoA requires transfer of an
AMP moiety to acetate
• Hydrolysis of pyrophosphate (PPi) product
drives reaction to completion
Fig 10.12 Synthesis of acetyl CoA
(continued next slide)
Fig. 10.12 (continued)
10.8 Thioesters Have High Free
Energies of Hydrolysis
• Thioesters are energy-rich compounds (10.22)
• Acetyl CoA has a Go’ = -31 kJ mol-1 (10.23)
Succinyl CoA Energy Can Produce GTP
10.9 Reduced Coenzymes Conserve
Energy from Biological Oxidations
• Amino acids, monosaccharides and lipids are
oxidized in the catabolic pathways
• Oxidizing agent - accepts electrons, is reduced
• Reducing agent - loses electrons, is oxidized
• Oxidation of one molecule must be coupled with
the reduction of another molecule
Ared + Box
Aox + Bred
A. Free-Energy Change Is Related
to Reduction Potential
• The reduction potential of a reducing agent is
a measure of its thermodynamic reactivity
• The electromotive force is the measured
potential difference between two half-cells
• Reference half-cell reaction is for hydrogen:
2H+ + 2e-
H2
Fig 10.13 Diagram of an electrochemical cell
• Electrons flow
through external
circuit from Zn
electrode to the
Cu electrode
Standard reduction potentials and free energy
• Relationship between standard free-energy
change and the standard reduction potential:
Go’ = -nFEo’
n = # electrons transferred
F = Faraday constant (96.48 kJ V-1)
Eo’ = Eo’electron acceptor - Eo’electron donor
Actual reduction potentials (E)
• Under biological conditions, reactants are not
present at standard concentrations of 1 M
• Actual reduction potential (E) is dependent
upon the concentrations of reactants and
products
E = Eo’ - (RT/nF) ln ([Aox][Bred] / [Ared][Box] )
B. Electron Transfer from NADH
Provides Free Energy
• Most NADH formed in metabolic reactions in
aerobic cells is oxidized by the respiratory
electron-transport chain
• Energy used to produce ATP from ADP, Pi
• Half-reaction for overall oxidation of NADH:
NAD+ + 2H+ + 2e-
NADH + H+ (Eo’ = -0.32V)
10.10 Experimental Methods
for Studying Metabolism
• Add labeled substrate to tissues, cells, and
follow emergence of intermediates
• Use sensitive isotopic tracers (3H, 14C etc)
• Verify pathway steps in vitro by using isolated
enzymes and substrates
• Use metabolic inhibitors to identify individual
steps and sequence of enzymes in a pathway