Chapter 9 (Continued)
The Pentose Phosphate Pathway
§
The Hexose
Monophosphate Shunt
§
Can be used at the same time as the glycolytic pathway
or the ED.
§
A pathway that is, in its entirety, is seldom used by anaerobic microorganisms,
but can be operated both aerobically and anerobically.
§
Aerobic organisms can use the complete pathway to
produce Ribulose 5-phosphate:
o
which is used in the biosynthesis of nucleic acids and to produce reduced NADP
§
The NADPH produced
by HMP is used for anabolic – biosynthesis,
rather than catabolic pathways.
§
In addition to the 5 carbon sugars, HMP also produces 4-C
sugars - erythrose 4-phosphate:
o
Needed for synthesis
of aromatic amino acids.
§
If the microorganism is using a 5 carbon sugar as its
sole carbon source, the HMP can serve as a source of 6-carbon sugars:
o
glucose is required
for peptidoglycan synthesis
§
Thus the HMP has both catabolic and anabolic functions,
this is a good example of an AMPHIBOLIC
pathway.
§
Two unique
enzymes of this pathway:
1. Transketolase: catalyzes the transfer of a 2-carbon ketol group
2. Transaldolase: transfers a three-carbon
group from sedoheptulose 7
phosphate to
glyceraldehydes 3 P.
§ Glucose 6Pà Fructose 6P + G3P
§ G3P enter 3 Carbon stage of Glycolytic pathway
§ Then converted:
G3P à ATP + Pyruvate.
Production
of NADPH, which can be converted to
NADH yielding ATP when oxidized by
electrons
transport chain.
Entner-Douderoff
Pathway
§
Some microorganisms lack the embden meyerhoff pathway
for the metabolism of glucose. These
organisms use a different pathway, which was discovered by Entner and
Doudoroff.
§
The initial substrate in the ED and EMP pathway is glucose, but the two pathways differ in
the final electron acceptor molecule and
the total energy yield.
§
The cell expends one ATP to make glucose 6-phosphate and
then converts it to 6 phospho-gluconate using NADP+ to produce NADPH.
§
6 phospho-gluconate is dehydrated to form a 6 carbon
unit the key intermediate in this pathway, 2 keto-
3-deoxy-6-phosphogluconate
or KDPG.
§
KDPG is then
cleaved by an aldolase (KDPG aldolase) to yield pyruvate and G3P.
§
G3P à to pyruvate
with the production of ATP and NADH (Same as the bottom part of the glycolyitc
pathway).
§
If ED pathway each glucose metabolized to pyruvate the
yield of energy is:
o
1 ATP
o
1 NADPH
o
1 NADH
Calculate:
used ATP to phosphate glucose
G6P
to 6-Pgluconate (make NADPH)
§
The ED is pathway is not used by all microorganisms,
it is frequently used by aerobic
microorganisms, which rely on
respiration for the majority of their ATP synthesis.
§
Anaerobic microorganisms, as a rule will not use ED,
but will use HMP because they can generate
more
ATP.
§
Mostly seen in the gram negative bacteria. Very few gram positive have with the
exception of Enterococcus faecalis.
Fermentaion
§
Pyruvic acid is readily available from the breakdown
of glucose in a variety of metabolic pathways.
§
Many cells use pyruvate,
or molecules derived from pyruvate as a terminal electron acceptor to
produce waste products that are exported out of the cell.
Lactic acid fermentation
NADH NAD+
\ /
Pyruvate ————————————> Lactate
Reduction
This fermentation pathway is found in many bacteria
and is
exploited in the production of yogurt, cheese,
buttermilk,
sourcream, and that large clump that forms in your
milk
carton.
Two Groups of
Lactic Acid Fermenters:
1. Homolactic Fermenters:
§
Organisms which produce only lactic acid
2. Heterolactic
Fermenters:
§
organisms which can produce other products in
addition to lactate ie EtOH and/or Lactate,
CO2
Alcoholic
fermentation:
§
Alcohol is produced by a two step reaction process.
o
First pyruvate is decarboxylated to acetaldehyde
which
is in turn reduced to Ethanol with NADH as the electron donor.
§
Fungi, some bacteria, algae, and protozoa.
Formic acid
fermentation:
§
Many bacteria (Enterobacteriaceae) are able to split
the 3 carbon pyruvate into formic acid (1-c) and a variety of other metabolic
products.
§
The pathways used to produce formate is a very useful
tool in identifying the enteric bacteria.
There are essentially two routes to format...
Mixed Acid Fermentation:
§
Ethanol is produced
and secreted along with a complex mixture of other fermentation products,
particularly acetic, lactaic, succinic
and formic acids.
§
If the organism possesses the enzyme complex formic hydrogenylase, it will convert
formic acid into CO2 and H2 gas.
§
Mixed acid fermentation produces considerably more acid than the other fermentation pathways,
and consequently results in a
significant change in pH
§
This fermentation pattern is seen with Escherichia,
Salmonella, Proteus, and other genera.
Butanediol fermentation:
§
Pyruvate is converted to acetoin, which is
subsequently reduced to 2,3-butanediol
§
This pathway produces
a large amount of alcohol, CO2
and H2, and very little acid via the mixed acid pathways.
§
It is important to emphasize that the cells are not
committed to a single pathway... some of the pyruvate in still being converted
to formate via alternate routes, but the
vast majority is being converted to butanediol
§
Butanediol fermentation is a characteristic property
of bacteria belonging to Enterobacter,
Serratia, Erwinia genera, and some species of Bacillus.
§
The butanediol fermenters produce neutral end products while mixed acid fermenters produce a
significant amount of acid end-products
(4 times) which can drop the pH lower than 4.4.
§
The acid end products can easily be identified by
adding a pH indicator to the growth medium, (ie methyl red (red @ acid)) but
the neutral end products will not react
§
The voges proskauer test is designed to identify butanediol fermenters by detecting acetion,
the precursor of butanediol.
The
Tricarboxylic Acid Cycle – Citric Acid Cycle –
(Krebs
Cycle):
In this cycle:
§
Energy is released from the breakdown of glucose to
pyruvate, much more energy is released when pyruvate is degraded aerobically to
CO2 .
§
TCA is a 3
stage Catabolic process
1. Attachment of a acetyl group to the acetyl
carrier, oxaloacetate to form citrate.
2. Begins with citrate and end in the formation
of succinly CO-A: acetyl carrier portion of
citrate loses two carbons when
oxidized to CO2.
3. Convert succinyl-Co back to oxaloacetate,
the acetyl carrier, so that it can
pick up another acetyl group.
The TCA cycle consists of
a set of 8 enzymes that further
break down the two acetyl Co-A
molecules into 4 CO2
TCA figure:
§
2 (3-carbon)
pyruvate molecules are broken down into
2 (2 carbon) acetyl coenzyme A molecules (acetyl-CoA)
(an energy-rich molecule) and 2 CO2 molecules by
§
Pyruvate Dehydrogenase(multi-enzyme
system)
an
enzyme reaction which links the Embden Meyerhoff pathway and the Citric Acid Cycle.
Isocitrate oxidixed and decarboxylated (2x) à alpha ketoglutarate.
o
2 NADHs
o
2 Carbons are lost as CO2 maintaining the balance because 2
carbons were added at the beginning.
Succiny CoA à Succinate
o
substrate level phosphorlyation, where GTP (high
energy molecule equivalent to ATP)is formed
Succinate à Fumarate (Four carbon stage)
o
1 FADH2
o
1 NADH
TCA cycle generates:
o
2 CO2s
o
3 NADHs
o
1 FADH2
o
1 GTP
For each acetyl CoA molecule oxidized.
Electron Transport Chain
§
The reduction of O2 occurs in the cell
membrane and is mediated by a series of proteins which are collectively called
the electron transport chain.
§
The electron transport chain consists of:
o
Cytochromes:a series of
heme containing proteins that transport electrons.
o
a series of nonheme iron sulfur proteins and quinones
§
The electron transport system is essentially a proton
pump which establishes a proton gradient across the cell membrane
§
As electrons flow through the ETS, they are
periodically moved from the inside to the outside of the cell membrane
o
Electron Transport Chain carriers are on the inside of
the inner membrane of the mitochondrion
or the bacterial plasma membrane
§
This translocation of proton drops the pH of the
periplasm to ~ 5.5 while the cytoplasm (inside) remains at ~8.5, a difference
of 3 pH units or 1000X concentration difference. This charge differential represents potential energy which is
stored up in the proton gradient.
§
This energy is referred to as the proton motive force
Proton Motive Force:
§
The force arising from a gradient of protons and a
membrane potential that is thought to power ATP synthesis and other processes.
Since the membrane is impermeable to protons, the only
way that these protons can re-enter the cell is through a transport protein.
o
ATP synthase is a protein
complex which forms a channel that will let protons pass through the membrane.
o
As the protons push through the channel, there is energy
used to convert ADP and Pi to ATP
o
Oxidative Phosphorylation or chemi-osmotic phosphorylation (Peter Mitchell
1961) is when the energy from electeon
transport is used to make ATP.
o
Substrate
level phosphorylation refers to the
synthesis of ATP by phosphoryl group transfer directly catalyzed by catabolic enzymes.
The second way that a cell can store energy is by
maintaining a proton (or charge) gradient across the cell membrane.
The phopholipid bilayer that composes the cell
membrane is impermeable to protons. The
cell uses a set of transport proteins to pump protons across the membrane, out
of the cell
This creates a proton gradient across the membrane,
High proton outside...low proton inside.
(the proton gradient is a reservoir of potential
energy that can be harnessed in a controlled fashion to generate high energy
bonds in the form of ATP)
The cells can then allow the protons to re-enter the
cell in a controlled fashion, and the energy derived from this movement can be
used to do work.
This is analogous to water stored behind a dam Respiration
In respiration, the terminal electron (or H) acceptor molecules are generally inorganic. There are some exceptions, such as with fumarate and trimethylamine oxide)
In aerobic respiration, molecular oxygen is used as the terminal electron acceptor
In anaerobic respiration, comp[ounds such as NO3-, SO3- (sulfate) and other compound substitute for oxygen as the final H acceptor molecule
The equasion for aerobic respiration can be expressed as follows: