Chapter
8 - Metabolism of Microbes
What
is metabolism? All the biochemical reactions that take place in a
cell.
The
Model: Metabolism
leading to the synthesis of a new microbial cell has 3 requirements:
1. Raw
Materials - nutrients composed of carbon (carbohydrates, proteins, etc.)
2. Driving
Force
a.
energy
- (ATP - adenosine triphosphate) - Some chemical
produce ATP; some chemical reactions use ATP.
b. reducing
power
- Many biochemical reactions involve oxidation (removal of electrons from a
compound) & reduction (addition of electrons to a compound; E. coli
stores electrons in coenzymes called NAD (nicotinamide adenine dinucleotide) & NADP
(nicotinamide adenine dinucleotide phosphate). These
compounds capture electrons in the form of hydrogen atoms from compounds that are being
oxidized, thus forming NADH & NADPH). NAD & NADP stores the cell's reducing power. Bacteria will then use this reducing power to
build its cellular components (it will reduce other compounds in this process).
Hint: O.I.L. = oxidation is loss; R.I.G. = reduction is
gain
3. A
Plan - contained in the cell's DNA; this is “the genetic code.”
A. Flow
of Materials - Sequential Steps:
1. Entry
Mechanisms
- Raw materials from the environment are transported into the cells by various mechanisms.
2. Catabolic
Reactions
- reactions in which raw materials are broken down
into smaller molecules (precursor molecules);
there are 12 precursor molecules that are required to synthesize building blocks (amino acids, monosaccharides,
nucleotides, fatty acids) that will build a new cell.
3. Anabolic
Reactions
- reactions that build up larger molecules from
smaller ones:
a.) biosynthesis
- 12 precursor molecules are put together to produce building blocks.
b.) polymerization
- building blocks are joined together to form macromolecules
(proteins, nucleic acids, lipids, polysaccharides, peptidoglycan); ex. amino acids are put
together to form a protein, nucleotides are put together to form DNA.
c.) assembly
- macromolecules
are assembled into biological structures; ex. peptidoglycan forms a cell wall.
raw
materials à
precursor molecules à
buliding blocks àmacromolecules
B. Driving
Force - Where is ATP & Reducing Power Produced & Used in the Above Reactions?
1. Entry
Mechanisms
- Many materials that move into the cell are moving from low to high concentration; this
requires ATP; remember the bacterial is usually hypertonic to its environment.
2. Catabolic
Reactions
- In general, the catabolic reactions transform raw materials into precursor molecules,
reducing power, & ATP.
3. Anabolic
Reactions
- In general, these reactions use precursor molecules, ATP, & reducing power.
III. Aerobic
Metabolism (We'll use E. coli as the example)
(Aerobic
means that this type of metabolism requires oxygen)
A. Catabolic
Reactions
- Supply the cell with the 12 precursor molecules, reducing power (NADH & NADPH),
& ATP.
1. Precursor
Molecules -
A minimum of 3 different pathways are required to produce all 12 precursor molecules: (We’ll look at these later in this handout)
a. glycolysis
- produces 6
b. tricarboxylic
acid (TCA) cycle - produces 4
c. pentose
phosphate pathway - produces 2 (we
won’t discuss this one)
2. Reducing
Power
- Many steps in a catabolic pathway involve oxidation (loss of electrons)/reduction
(gain of electrons) reactions. The electrons
(along with hydrogens) are usually transferred from the molecule being oxidized to the
coenzymes NAD or NADP, to form NADH & NADPH. NADH & NADPH are used to reduce other
molecules or are used to generate new ATP molecules.
3. ATP
a.
Stored
Energy
- The principal compound that stores chemical energy in all cells is ATP (adenosine
triphosphate). The energy-storage
capabilities of ATP depend on the 2 bonds that join the 3 phosphate groups in the ATP
molecule. These bonds are among the most highly
reactive bonds found in biochemicals. They
are called high-energy bonds. The
phosphate groups that are joined in ATP by high-energy bonds are readily donated to other
compounds. These compounds that receive a
phosphate group from ATP are termed phosphorylated compounds. After receiving a
high-energy phosphate, phosphorylated
b.
ATP Formation from ADP [most of the ATP is made by chemiosmosis]
1.) Chemiosmosis
& the Electron Transport System-
The electron transport system is made up of a series or chain of compounds. Some of these compounds are hydrogen-carriers and
some are electron-carriers. NADPH/NADH transfers hydrogen atoms to a hydrogen-carrier in
the chain. This hydrogen-carrier then passes
hydrogen electrons to an electron-carrier in the chain; the hydrogen ions are pumped out
of the cell (or out of the inner mitochondrial compartment, if you're talking about
eukaryotes). Each compound in the chain will
then alternately accept & then release hydrogen electrons; as electrons are passed
along the chain they drop to lower and lower energy levels.
At the end of the chain, electrons are accepted by oxygen to form water. (The final electron acceptor is oxygen!! This is why this process is called aerobic respiration!).
ATPase is an enzyme located in the cell
membrane of prokaryotes; in eukaryotes it's located in the inner mitochondrial membrane. In E. coli, the pumping of hydrogen ions (H+)
out of the cell creates a H+ concentration gradient & an electrical
gradient across the cell membrane (there are more H+ outside the cell than
inside). H+ then flows back into
the cell (H+ ions want to move from a high concentration to a low
concentration). The ions move through the
membrane-bound ATPase, converting ADP to ATP. This
process is called chemiosmosis.
4. Catabolic
Pathways (breaking
down) - Chemical reaction sequences that transform raw materials into the 12 precursor
molecules, store energy in the form of ATP, & store reducing power in the form of
NADPH/NADH. Central metabolism (described below) begins with
the monosaccharide glucose. Most
organisms have other catabolic pathways, which use substrates other than glucose (ex.
humans can use protein & lipids, if glucose concentrations are low). These pathways all eventually feed into central
metabolism at various points.
a.
Glycolysis (=
"glucose splitting") - This pathway begins with glucose (a 6 carbon sugar) and
ends with 2 pyruvic acids or pyruvates (3 carbon molecules). The initial reactions in glycolysis require ATP. Later reactions in glycolysis produce a small
amount of ATP. A small amount of NADH is also
produced, which will result in the production of more ATP in the electron transport
system. The main function of glycolysis is to
split glucose.
b. Kreb's
Cycle -
Some of the pyruvate formed by glycolysis is used in biosynthesis, the rest is oxidized to
another precursor molecule, acetyl CoA (coenzyme A).
Acetyl-CoA then enters the Kreb's Cycle by combining with a precursor molecule
oxaloacetate to form citrate. In a series of
6 reactions, carbon dioxide is released & oxaloacetate is regenerated (this is where
most of the carbon dioxide you exhale comes from!). In
this cycle, only a small amount of ATP is formed (called substrate level phosphorylation). However, considerable reducing power is stored in
the form of NADH, NADPH, & FADH2 (flavine adenine dinucleotide), which are
used to produce ATP in the electron transport system.
c. Pentose
Phosphate Pathway - we won’t discuss.
5. Anabolic
Pathways (building
up) - use precursor molecules, ATP, &
reducing power produced in above catabolic reactions.
a.
Biosynthesis
- E. coli converts precursor molecules produced in catabolism into building blocks
(amino acids, monosaccharides, nucleotides, fatty acids) of macromolecules (proteins,
nucleic acids, lipids, polysaccharides, peptidoglycan).
Organisms that can't make a given building block grow only if that molecule is
provided in the medium (or diet). Ex. E. coli can make all 20 amino acids. Humans are unable to make 9 of the 20, so these
nutrients must come from our diets.
b. Polymerization
- In these reactions, building blocks produced in biosynthesis are joined to form
macromolecules. The major cellular
polymerization reactions are DNA replication, RNA synthesis, protein synthesis, &
polysaccharide & peptidoglycan synthesis. For most macromolecules, building blocks must
be joined in a specific order. For
ex., amino acids must be arranged in the proper order to produce the right proteins. If you change the amino acid sequence, you change
the protein’s structure and therefore change its function.
Some
polymerization is determined directly by the organism's DNA (ex. DNA replication,
RNA synthesis, protein synthesis). Other
reactions are indirectly determined by DNA (ex. polysaccharide & peptidoglycan
synthesis). In the latter, building blocks
are ordered by the enzymes that catalyze the polymerization reactions. Because enzymes are proteins & protein
structure is determined by DNA, the reactions the enzymes catalyze are indirectly
determined by DNA.
c.
Assembly
- Assembly of macromolecules into cellular structures (ex. flagella, ribosomes, cell wall)
may occur spontaneously (self-assembly) or as a result of reactions catalyzed by
enzymes.
Anaerobic
metabolism
allows cells to grow in the absence of oxygen.
Strict
anaerobes
are capable of only anaerobic metabolism.
Facultative
anaerobes
are capable of both aerobic & anaerobic metabolism.
(ex. E. coli)
A. Anaerobic Respiration
- Involves an electron transport chain, but uses a compound other than oxygen as the final
electron acceptor, allowing the cell to generate ATP by chemiosmosis. Compounds that can be used as final electron
acceptors include sulfate & nitrate.
1. Nitrate
users:
These organisms, including E. coli, play a role
in the nitrogen cycle (removing nitrogen from terrestrial & aquatic environments &
returning it to the atmosphere). Some
microbes reduce nitrate to nitrite. Some
microbes reduce nitrite further to nitrogen gas. We’ll
see this in lab!
2. Sulfate
users:
These organisms, called sulfur reducers, play a role in the sulfur cycle. Sulfate is reduced to hydrogen sulfide gas. Sulfate reducers typically grow in marine &
river mud flats, giving these environments a rotten-egg odor & turning the mud black. We’ll see this in lab!
B. Fermentation
- This type of anaerobic metabolism uses no electron transport chain.
Some differences between fermentation & respiration (aerobic or anaerobic):
1. Fermentation
generates fewer ATP per molecule of substrate. (ex. E. coli can produce 38 ATP/molecule of glucose by
aerobic respiration, but only 3 ATP by fermentation.)
2. Because
many molecules of substrate must be metabolized to supply a cell's ATP requirements, the
substrate must be in abundance in order for the microbe to grow.
3. Sugars
are usually the only substrate that can be used in fermentation.
1. Lactic
Acid Fermentation
- This type of fermentation is carried out by lactic acid bacteria, the microbes that
cause milk to sour (used to produce yogurt & buttermilk). Muscle tissue of animals also carries out lactic
acid fermentation, when deprived of oxygen (during strenuous exercise - this is what makes
your muscles sore). Steps:
1. Glycolysis
- Glucose is metabolized to produce 2 molecules of pyruvic acid (a little ATP & NADH
is also produced).
2. Pyruvic
Acid Oxidation - pyruvic acid is oxidized to lactic acid using NADH, thus using up the
reducing power stored in glycolysis.
Parts
of the Kreb's cycle & Pentose Phosphate pathway are used to help generate the 12
precursor metabolites; they cannot all be formed in fermentation.
2. Alcoholic Fermentation
This
type of fermentation is typical of yeast, a type of fungi.
In this pathway, pyruvic acid is converted to carbon dioxide and ethanol.
glucose à
pyruvic acid à
carbon dioxide + ethanol
Microbes are classified according to nutritional class, which depends on 2 factors:
1.) How
it generates ATP & reducing power: chemo- (from oxidation of inorganic compounds like
sulfur nitrite, ammonia, iron) or photo-
(sun)
2.) The
source of carbon atoms it uses to make precursor metabolites: auto-
(obtain carbon from carbon dioxide) or hetero-
(obtain carbon from organic compounds like carbs, proteins, lipids, etc.)
Can
have combinations of all of these:
chemoautotrophs,
chemoheterotrophs, photoautotrophs, photoheterotrophs.