Transcript Document

Basic Concepts of Cellular
Metabolism and Bioenergetics
Intermediary Metabolism
The Chemistry of Metabolism
Concepts of Bioenergetics
Experimental Study of Metabolism
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Metabolism
Metabolism
The summation of all chemical reactions
in an organism.
Metabolic differences are best studied by
dividing all life into two categories.
Autotrophs - organisms that use atmospheric
CO2 as their sole source of carbon.
Heterotrophs - life forms that obtain energy by
ingesting complex carbon compounds .
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Intermediary metabolism
Metabolism relies on thousands of sequential
enzymatically controlled reactions.
Intermediary metabolism. Products from one
reaction often become the reactant for the
next - metabolites.
Pathway. A series of reactions with a specific
purpose.
Linear - Glycolysis
Branched - Amino acid biosynthesis
Cyclic - Citric Acid Cycle
Spiral - Fatty acid degradation
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Intermediary metabolism
Two paths of metabolism:
Catabolism
Degradation path. Complex organic
molecules are degraded to simpler
species. Production of energy.
Anabolism
Construction path. Biosynthesis of more
complex organic compounds. Requires
energy.
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Energy
Energy, ATP and the
movement of phosphate
phosphoenolpyruvate
P
1,3-bisphosphoglycerate
P
creatine phosphate
P
ADP
ATP
glucose-1-phosphate
P
fructose-6-phosphate
P
glucose-6-phosphate
P
ADP
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ATP
ATP adenosine triphosphate
a nucleotide composed of three basic units.
adenine
phosphate chain
O
-
O
P
O-
O
O
P
O-
NH2
O
O
N
P
O
O-
CH2
N
O
OH
N
N
OH
ribose
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ATP and ADP
NH 2
O-
O
P O
O-
ADP
O
P O
O- CH 2
N
N
O
N
N
Energy is
released when
it is removed.
OH
OH
NH 2
O
O- P O
O-
O
P O
O-
O
P O
O- CH 2
N
N
O
ATP
OH
OH
It takes energy
to put on the
third phosphate.
N
N
ADP - ATP
conversions act
as a major
method of
transferring
energy.
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Catabolic stages of metabolism
Stage I
Breakdown of macromolecules into their
building blocks. No useful energy.
Stage II
Oxidation of Stage I products to acetyl CoA.
Limited energy production.
Stage III
Oxidation of acetyl CoA to CO2 and H2O and
energy.
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Overview of
catabolic processes
Proteins
Fats
Carbohydrates
Stage 1
Amino acids
Fatty acids
Simple Sugars
Glycolysis
Stage 2
Pyruvate
ATP
Acetyl CoA
Citric acid cycle
Stage 3
Oxidative phosphorylation
ATP
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Overview of catabolic metabolism
protein
polysaccharides
ADP + Pi
lipids
ADP + Pi
ATP
ATP
ADP + Pi
ATP
hexoses
pentoses
amino acids
ADP + Pi
ADP + Pi
ATP
ATP
fatty acids
ADP + Pi
ADP + Pi
ATP
ATP
pyruvate
urea
ADP + Pi
acetyl-CoA
urea
cycle
CO2
ATP
citric acid
cycle
e-
O2
electron transport
chain
oxidative
phosphorylation
ATP
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Stage one
Hydrolysis of food into smaller subunits.
Handled by
the digestive
system.
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Stage one
Salivary glands
Secrete amylase - digests starch.
Stomach
Secretes HCl - denatures protein and
pepsin.
Pancreas
Secretes proteolytic enzymes and lipases
- degrades proteins and fats.
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Stage one
Liver and gallbladder
Deliver bile salts.
- emulsify fat globules - easier to digest.
Small intestine
Further 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 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-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-Reduction
When an atom or group is oxidized, some
other species must accept the electrons.
Many reactions are coupled to the
coenzyme pairs.
NAD+ / NADH
NADP+ / NADPH
FAD / FADH2
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Coenzymes used
in metabolism
NAD+
NADH
Oxidized 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 used
in metabolism
reactive
site
O
O
O
C
-
P
O
CH2
O
O
O
O
CH2
-
ribose
nicotinamide
N+
O
OH
P
NH2
OH
N
N
O
OH
NH2
N
adenine
N
OH
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NAD+
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Coenzymes used
in metabolism
Example reactions of NAD+
General reaction
OH
R C H + NAD+
H
O
R C H + NADH + H +
Specific example - ethanol
CH3CH2OH + NAD+
alcohol
dehydrogenase
H
CH3C=O + NADH + H+
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Coenzymes used
in metabolism
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 used
in metabolism
FAD
O
H3C
N
H3C
Reactive site
is highlighted
NH
N
N
H
C
H
H
C
OH
H
C
OH
H
C
OH
H
C
H
O
riboflavin
NH2
N
O
O
P
O
O
-
ribose
CH2
N
O
OH
N
adenine
N
OH
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FAD
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Coenzymes used
in 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
||
CH3CH2CH2-C-S-CoA
O
||
CH3CH=CHC-S-CoA
FAD FADH2
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Group-Transfer
Reactions that involve moving a chemical
functional group.
Intermolecular. Transfer from one molecule
to another.
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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Group-Transfer
Another common group to transfer is acyl
group.
O
R-C
Coenzyme A (CoASH) will form a thioester
linkage to this group, making it more
active.
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Acetyl - coenzyme A
phosphorylated
ADP
pantothenate
unit
NH2
O
O H CH3
C-CH2-CH2-N-C-C-C-CH2
H HO CH3
H-N
CH2-CH2
S
CH3C
O
O
N
N
P O P O
O-
Sulfhydyl
group
O- CH 2
O
O
N
N
OH
O P OO-
acetate
This molecule serves as the carrier
for the small molecules from digestion.
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Acetyl - coenzyme A
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Hydrolysis
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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Hydrolysis
Carbohydrates
CH2 OH
O H
H
OH H
OH H
O
H
OH
H
CH2 OH
O
H
OH H
H
OH
+ H2O
H
OH
enzyme
CH2 OH
O H
H
OH H
OH H
H
OH HO
OH
H
CH2 OH
O
H
OH H
H
OH
H
OH
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Hydrolysis
Proteins
H O
H
| ||
|
H2N - C - C - N - C - COOH
|
| |
R
enzyme
H R’
+ water
H
|
H2NCCOOH +
|
R
H
|
H2NCCOOH
|
R’
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Hydrolysis
Fats
O
H
H C
O
O
H
C
O
R
H C
HO
R
OH
O
H C
O
C
O
R’ + 3 H2O
H C
OH
H C
O
C
R’’
H C
OH
H
C
H
+
HO
C
R’
O
HO
C
R’’
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Nonhydrolytic cleavage
A class of reactions where molecules are
split without the use of water.
Lyases - Enzymes that accomplish this task.
CH2OPO 32C O
HO C H
HO C H
HO C H
aldolase
CH2OPO 32C O
H
O
C
+ HO C H
CH2OH
CH2OPO 32-
CH2OPO 32-
fructose-1,6bisphosphate
dihydroxyacetone
phosphate
glyceraldehyde
3-phosphate
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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
H
O
C
H C OH
H C OH
HO C H
C OH
HO C H
H C OH
H C OH
H C OH
H C OH
CH 2OH
aldose
CH 2OH
cis-enediol
intermediate
CH2OH
C O
HO C H
H C OH
H C OH
CH 2OH
ketose
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Bond formation reactions
using energy
Category of biochemical bond formation
reactions. All require an energy source.
COOHO C H
NAD +
NADH + H +
H C COOH
H C H
COO-
isocitrate
DHase
COO-
COO-
C O
C O
H C COO-
H C H
H C H
H C H
COO-
COO-
oxalosuccinate
+ CO2
-ketoglutarate
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Concepts of bioenergetics
Standard free energy change - Go
The energy change occurring when a
reaction, under standard conditions,
proceeds from start to equilibrium.
Equilibrium
A + B
K’eq =
C + D
[C] [D]
[A] [B]
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Standard free energy changes
Go can be related to the equilibrium
expression by:
Go’ = -2.303 RT log K’eq
where
Go’
R
T
K’eq
standard free energy change
gas constant, 8.316 J/mol
temperature, kelvin
equilibrium constant
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Standard 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 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 Go’
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 ]
[ fructose-6-phosphate ]
= 1.33 M
= 0.67 M
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Experimental measurement of Go’
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 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 ATP
Using a coupled reaction, it is possible to
measure the Go’ for ATP.
Go’ kJ/mol
glucose + ATP
glucose-6-phosphate + ADP
glucose-6-phosphate + H2O
glucose + Pi
-16.7
-13.8
Sum: ATP + H2O
-30.5
ADP + Pi
This is a relatively large amount of useful
chemical energy.
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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
Whole organisms.
One can introduce radiolabeled materials
and measure any labeled waste products.
Tissue slices and cells.
These have been used to uncover
metabolic details. The citric acid cycle
was characterized using this approach.
Cell-free extracts.
Cells are homogenized in a buffer to
release cell components for study.
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