1 basic concepts of cellular metabolism and bioenergetics intermediary metabolism the chemistry of...
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Basic Concepts of Cellular Metabolism and Bioenergetics
Basic Concepts of Cellular Metabolism and Bioenergetics
Intermediary Metabolism
The Chemistry of Metabolism
Concepts of Bioenergetics
Experimental Study of Metabolism
Intermediary Metabolism
The Chemistry of Metabolism
Concepts of Bioenergetics
Experimental Study of Metabolism
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MetabolismMetabolism
MetabolismMetabolismThe summation of all chemical reactions in an organism.
Metabolic differences are best studied by dividing all life into two categories.
AutotrophsAutotrophs - organisms that use atmospheric CO2 as their sole source of carbon.
HeterotrophsHeterotrophs - life forms that obtain energy by ingesting complex carbon compounds .
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Intermediary metabolismIntermediary metabolism
Metabolism relies on thousands of sequential enzymatically controlled reactions.
Intermediary metabolism.Intermediary metabolism. Products from one reaction often become the reactant for the next - metabolites.
Pathway.Pathway. A series of reactions with a specific purpose. Linear - GlycolysisLinear - Glycolysis
Branched - Amino acid Branched - Amino acid biosynthesisbiosynthesis
Cyclic - Citric Acid CycleCyclic - Citric Acid Cycle
Spiral - Fatty acid degradationSpiral - Fatty acid degradation
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Intermediary metabolismIntermediary metabolism
Two paths of metabolism:
CatabolismCatabolismDegradation path. Complex organic molecules are degraded to simpler species. Production of energy.
AnabolismAnabolismConstruction path. Biosynthesis of more complex organic compounds. Requires energy.
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Energy, ATP and themovement of phosphate
Energy, ATP and themovement of phosphate
ATP
ADP
ADP
phosphoenolpyruvatephosphoenolpyruvate
1,3-bisphosphoglycerate1,3-bisphosphoglycerate
creatine phosphatecreatine phosphate
glucose-1-phosphateglucose-1-phosphate
fructose-6-phosphatefructose-6-phosphate
glucose-6-phosphateglucose-6-phosphate
P
En
erg
y
P
P
P
P
P
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ATPATP
ATPATP adenosine triphosphateadenosine triphosphatea nucleotide composed of three basic units.
adenine
phosphate chain
ribose
CH2 O
OH OH
N
NN
N
NH2
OPOPOPO-
O
O-
O
O-
O
O-
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ATP and ADPATP and ADP
CH 2 O
OH OH
N
NN
N
NH 2
OPOPO-
O
O-
O
O-
ADP
CH 2 O
OH OH
N
NN
N
NH 2
OPOPOPO-O
O-
O
O-
O
O-
ATP
It takes energyto put on thethird phosphate.
Energy isreleased whenit is removed.
ADP - ATP conversions actas a major method oftransferringenergy.
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Catabolic stages of metabolismCatabolic stages of metabolism
Stage IStage IBreakdown of macromolecules into their building blocks. No useful energy.
Stage IIStage IIOxidation of Stage I products to acetyl CoA. Limited energy production.
Stage IIIStage IIIOxidation of acetyl CoA to CO2 and H2O and energy.
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Overview of catabolic processes
Overview of catabolic processes
CarbohydratesCarbohydrates FatsFatsProteinsProteins
Simple SugarsSimple Sugars Fatty acidsFatty acidsAmino acidsAmino acids
PyruvatePyruvate
Acetyl CoAAcetyl CoA
Oxidative phosphorylationOxidative phosphorylation
ATP
ATP
Citric acid cycleCitric acid cycle
Stage 1
Stage 2
Stage 3
GlycolysisGlycolysis
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Overview of catabolic metabolismOverview of catabolic metabolism
acetyl-CoA
pyruvate ATP
ADP + Pi
polysaccharides
hexosespentoses
ADP + Pi
ATP
ADP + Pi
ATP
ADP + Pi
ATPATP
ADP + Pi
ADP + Pi
ATP
lipids
fatty acids
ATP
ADP + Pi
protein
amino acids
citric acidcycle
ureacycle ATP
ADP + Pi
urea
CO2
electron transportchain
oxidative phosphorylation
O2
ATP
e-
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Stage oneStage one
Hydrolysis of food into smaller subunits.
Handled bythe digestivesystem.
Handled bythe digestivesystem.
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Stage oneStage one
Salivary glandsSalivary glandsSecrete amylase - digests starch.
StomachStomachSecretes HCl - denatures protein and pepsin.
PancreasPancreasSecretes proteolytic enzymes and lipases - degrades proteins and fats.
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Stage oneStage one
Liver and gallbladderLiver and gallbladderDeliver bile salts.- emulsify fat globules - easier to digest.
Small intestineSmall intestineFurther degradation.Produces amino acids, hexose sugars, fatty acids and glycerol.Moves materials into blood for transport to cells.
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The chemistry of metabolismThe chemistry of metabolism
Six categories of biochemical reactions have been identified.
• Oxidation-reduction
• Group-transfer
• Hydrolysis
• Nonhydrolytic cleavage
• Isomerization and rearrangement
• Bond formation reactions using energy from ATP
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Oxidation-ReductionOxidation-Reduction
Most common of all metabolic reactions. • There are always two reactant molecules.
• They are readily identified by the transfer of hydrogen atoms.
• Enzymes involved in these reactions are oxidoreductases (dehydrogenases).
AH2 + B A + BH2
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Oxidation-ReductionOxidation-Reduction
When an atom or group is oxidized, some other species must accept the electrons.
Many reactions are coupled to the coenzyme pairs.
NADNAD++ / NADH / NADH
NADPNADP++ / NADPH / NADPH
FAD / FADHFAD / FADH22
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Coenzymes usedin metabolism
Coenzymes usedin metabolism
NADNAD++ NADH NADHOxidized form Reduced form of nicotinamide adenine dinucleotide.
• Used in REDOX reactions.
• It is a derivative of ADP and the vitamin nicotinamide.
• The reactive site is located on the nicotinamide portion of NAD+.
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Coenzymes usedin metabolism
Coenzymes usedin metabolism
OCH2
OH OH
OPO
O-
O
OCH2
OH OH
OPO
O-N
NN
N
NH2
N+
C
O
NH2
reactivesite
nicotinamide
adenine
ribose
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NAD+NAD+
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Coenzymes usedin metabolism
Coenzymes usedin metabolism
Example reactions of NAD+
General reactionGeneral reaction
Specific example - ethanolSpecific example - ethanol
CH3CH2OH + NAD+
H CH3C=O + NADH + H+
R COHH
HHH + NAD+ R C
OH + NADHH + HH++
alcoholdehydrogenase
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Coenzymes usedin metabolism
Coenzymes usedin metabolism
FAD - flavin adenine dinucleotide.FAD - flavin adenine dinucleotide.
Another major electron carrier used in metabolism.
It involves a two electron transfer so it picks up two hydrogen.
FAD FADH2
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Coenzymes usedin metabolism
Coenzymes usedin metabolism
FAD
OCH2
OH OH
OPO
O-N
NN
N
NH2
O
C HH
CH OH
CH OH
CH OH
CH H
N
NH3C
H3C N
NH
O
O
ribose
adenine
riboflavin
Reactive siteis highlighted
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FADFAD
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Coenzymes usedin metabolism
Coenzymes usedin metabolism
FAD typically reacts with different substrates than NAD+.
FAD is often involved in oxidation reactions in which a -CH2 - CH2 - portion is oxidized to a double bond.
O O
|| ||
CH3CH2CH2-C-S-CoA CH3CH=CHC-S-CoAFAD FADH2
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Group-TransferGroup-Transfer
Reactions that involve moving a chemical functional group.
Intermolecular.Intermolecular. Transfer from one molecule to another.
Intramolecular.Intramolecular. Movement from one location to another on the same molecule.
Phosphate is one of the most important groups that is transferred.
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Another common group to transfer is acyl group.
Coenzyme A (CoASH) will form a thioester linkage to this group, making it more active.
Group-TransferGroup-Transfer
R - C
O
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Acetyl - coenzyme AAcetyl - coenzyme A
This molecule serves as the carrier
for the small molecules from digestion.
phosphorylated ADP
pantothenateunit
acetate
CH2 O
O OH
N
NN
N
NH2
OPOPO
O-
O
O-
P O-O
O-
C-CH2-CH2-N-C-C-C-CH2
OO H
CH3HO
CH3
HH-N
CH2-CH2
S
CH3C O
Sulfhydylgroup
O
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Acetyl - coenzyme AAcetyl - coenzyme A
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HydrolysisHydrolysis
Water is used to split a single molecule into two separate molecules.
Most common types of bonds to split
• Esters - fats
• Amides - proteins
• Glycosidic - carbohydrates
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HydrolysisHydrolysis
CarbohydratesCarbohydrates
OOH
H
H
H
H
OH
CH 2 OH
H
OH
OH OH
HH
H
OH
CH 2 OH
H
OH
O
OOH
H
H
H
H
OH
CH 2 OH
H
OH
OH OH
HH
H
OH
CH 2 OH
H
OHOH HO
+ H2O
enzyme
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HydrolysisHydrolysis
ProteinsProteins
H |
H2NCCOOH
| R
+
H |
H2NCCOOH
| R’
H O | ||
H2N - C - C -
| R
H |N - C - COOH | |H R’ enzyme
+ water
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HydrolysisHydrolysis
FatsFats
O C R
O
C
H
H
O C R’
O
CH
O C R’’
O
C
H
H
OHC
H
H
OHCH
OHC
H
H
C R’’
O
HO
++ 3 H2OC R’
O
HO
C R
O
HO
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Nonhydrolytic cleavageNonhydrolytic cleavage
A class of reactions where molecules are split without the use of water.
LyasesLyases - Enzymes that accomplish this task.
CH2OPO32-
C O
C
C
C
CH2OPO32-
H
H
H
HO
HO
HO
CH2OPO32-
C O
CH2OH
C
C
CH2OPO32-
OH
HO H+aldolase
fructose-1,6- dihydroxyacetone glyceraldehydebisphosphate phosphate 3-phosphate
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Isomerization and rearrangement
Isomerization and rearrangement
This category involves two kinds of chemical transformations:
• Intermolecular hydrogen atom shifts to the location of a double bond. Most prominent example is the aldose-ketose isomerization.
• Intramolecular rearrangements of functional groups. These are rare.
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Isomerization and rearrangement
Isomerization and rearrangement
C
C
C
C
C
CH 2 OH
OH
OH
H
OHH
HO
H
H
OH
C
C
C
C
C
CH 2 OH
OH
OH
H
OH
HO
H
H
C
C
C
C
CH2OH
CH 2 OH
OH
OH
H
O
HO
H
H
OHH
aldosecis-enediol
intermediate ketose
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Bond formation reactionsusing energy
Bond formation reactionsusing energy
Category of biochemical bond formation reactions. All require an energy source.
COO-
C
C
C
COO-
H
COOH
H
HO
H
H
COO-
C
C
C
COO-
O
COO-
H
H
H
COO-
C
C
C
COO-
O
H
H
H
H
+ CO2
NAD+ NADH + H+
isocitrate oxalosuccinate -ketoglutarate
DHase
spontaneous
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Concepts of bioenergeticsConcepts of bioenergetics
Standard free energy change - Standard free energy change - GGoo
The energy change occurring when a reaction, under standard conditions, proceeds from start to equilibrium.
EquilibriumEquilibriumA + B C + D
K’eq =[ C ] [ D ]
[ A ] [ B ]
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Standard free energy changesStandard free energy changes
Go can be related to the equilibrium expression by:
GGo’ o’ = -2.303 RT log K’= -2.303 RT log K’eqeq
whereGo’ standard free energy change
R gas constant, 8.316 J/mol T temperature, kelvinK’eq equilibrium constant
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Standard free energy changesStandard free energy changes
• These types of measurements can be made by mixing the reactants at 1 molar, 25oC and a pH of 7 in a test tube.
• Unfortunately, they do not agree well with the conditions of a living cell.
• They do provide an estimate for comparing energy requirements among the many reactions in a cell.
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Standard free energy changesStandard free energy changes
Go’ = 0 System at equilibrium, no release or requirement of energy.
Go’ < 0 Reaction releases energy as it approaches equilibrium.
Go’ > 0 Reaction requires that energy be added to proceed in the direction indicated.
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Experimental measurement of GGo’o’
Experimental measurement of GGo’o’
As an example, let’s determine Go’ for the isomerization of glucose-6-phosphate to fructose-6-phosphate.
To start, solutions are mixed that result in an initial concentration of one molar for each species at standard conditions.
At equilibrium we have:[ glucose-6-phosphate ] = 1.33 M[ fructose-6-phosphate ] = 0.67 M
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Experimental measurement of GGo’o’
Experimental measurement of GGo’o’
K’eq = 0.67 M / 1.33 M
= 0.50
Go’ = (-2.303)(8.315 J/mol)(298 K) log(0.5)
= +1718 J/mol = +1.7 kJ/mol
This indicates that energy is required for glucose-6-phosphate to be converted to fructose-6-phosphate -- it is not spontaneous.
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Energy from ATPEnergy from ATP
We can conduct a similar experiment using ATP and ADP:
ATP + H2O ADP + Pi
After mixing and allowing to reach equilibrium, we find that the concentration of ATP is too low to measure.
We can’t directly obtain Go’ but at least we know that it must be negative.
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Energy from ATPEnergy from ATP
Using a coupled reaction, it is possible to measure the Go’ for ATP.
Go’ kJ/mol
glucose + ATP glucose-6-phosphate + ADP -16.7glucose-6-phosphate + H2O glucose + Pi -13.8
Sum: ATP + H2O ADP + Pi -30.5
This is a relatively large amount of useful chemical energy.
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Experimental study of metabolism
Experimental study of metabolism
To understand a pathway, one must know all of the details of each step.
• Characterization of each enzyme and coenzyme.
• Identification of the chemical pathway, including the substrate, intermediates, products and types of reaction.
• Identification of molecules and conditions that regulate the overall rate of the pathway.
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Experimental study of metabolism
Experimental study of metabolism
Whole organisms.Whole organisms.One can introduce radiolabeled materials and measure any labeled waste products.
Tissue slices and cells.Tissue slices and cells.These have been used to uncover metabolic details. The citric acid cycle was characterized using this approach.
Cell-free extracts.Cell-free extracts.Cells are homogenized in a buffer to release cell components for study.