Lecture 4: Fermentation - Exam 4

Question 1

What is fermentation? Who does it?

Question 2

Where do the reactions occur?

Question 3

How is ATP made?

Question 4

What are the e- donors and acceptors?

Question 5

What is the problem with oxygen? How do aerobes deal with it? Why can’t anaerobes use it?

Answer 5

Oxygen is a strong oxidizing agent, making it an excellent electron donor. However, during the stepwise reduction of oxygen in respiration, toxic and highly reactive intermediates are produce:
-Reactive oxygen species (ROS)
-ROS attack iron dependent enzymes.
Aerobic bacteria have evolved systems designed to detoxify ROS
-Catalase
-Peroxidase
-Superoxide dismutase
Anaerobes do not possess these protective systems.
-Depending on aerotolerance, some anaerobes may have some degree of protection
-Microaerophilic bacteria typically possess superoxide dismutase.
-Strict anaerobes have no protection from ROS and cannot tolerate any oxygen.

Question 6

What are the two types of anaerobic growth?

Answer 6

Anaerobic respiration (using nitrate, sulfate, fumarate as the terminal e- acceptor) and fermentation.

Question 7

What is fermentation?

Answer 7

A pathway in which NADH (or other reduced e- carrier) is reoxidized by metabolites produced by the pathway.
-Redox reactions occur in the cytosol, not the membrane
-ATP is produced by substrate-level phosphorylation.
Fermentations are named after the major end products they generate.

Question 8

What is anaerobic respiration?

Answer 8

Similar to aerobic respiration in that electrons extracted from a fuel molecules are passed through an electron transport chain, driving ATP synthesis.
-Sulfate (SO4), nitrate (NO3) carbon dioxide (CO2) or other molecules are terminal electron acceptor of ETC.

Question 9

Archaea called methanogens use _________ as a terminal electron acceptor, producing _________ as a waste product.

Answer 9

Carbon dioxide or acetic acid ; methane
(Methanogens are found in soils, in the digestive systems or ruminants or termites and many other habitats)

Question 10

With fermentation, what kind of electron acceptor is not involved?

Answer 10

NO exogenous terminal electron acceptor is involved

Question 11

What is the only energy yielding step in fermentation?

Answer 11

Glycolysis, with one or two extra reactions tacked on at the end to recharge NAD+.
-Organic molecules serve as both electron donors and acceptors
-The molecule being metabolized is not completely oxidized

Question 12

Fermentation is not unique to _______.

Answer 12

Prokaryotes

Question 13

What are the benefits of fermentation?

Answer 13

Food products (beer, cheese, bread, sauerkrauat, kimchi, yogurt), bioalcohol, organic solvents

Question 14

Fermentation vs Respiration

Answer 14

Fermentation:
-Occurs in cytosol
-Electron movement is not coupled to proton extrusion
-An organic molecules serves as electron donor and acceptor
-ATP generated via substrate-level phosphorylation
Respiration:
-Occurs in cell membranes
-Electron movement is coupled to proton extrusion to generate PMF
-ATP generated using PMF via ATP synthase

Question 15

Fermentation:
NAD+ is reduced to NADH (glycolysis) then back to NAD+ when…?

Answer 15

It donates its electrons to electron sinks

Question 16

What is a major concern with fermentation?

Answer 16

Reoxidation of the reduced NADH.
Reoxidation of NADH is vital. Lack of NAD+/NADH balance can result in stoppage of biological reactions due to lack of electron acceptors.

Question 17

Fermentation does not involved exogenous electron acceptors (unlike respiration), so…?

Answer 17

The fermentation pathways themselves have to produce the electron acceptors that can be used to oxidize NADH.
-These electron acceptors are called electron sinks.

Question 18

Fermentation: Why are the electron acceptors produced by the fermentation pathways called electron sinks?

Answer 18

Because they dispose of the electrons removed during the reoxidation of NADH by being excreted from the cell, into the medium.
-Thus, fermentations are characterized by the excretion of large amounts of reduced organic compounds, like alcohols, lactate, organic acids.

Question 19

Describe ATP generated by substrate-level phosphorylation.

Answer 19

-Energy yields are very low compared to respiration.

Question 20

________ is often an intermediate of various fermentations.

Answer 20

Pyruvate

Question 21

Fermentation:
PMF is often generated by…?

Answer 21

Reversing ATP synthase (remember, ATP Synthase can make or break ATP).

Question 22

Describe the basic metabolic adaptation to anaerobiosis.

Answer 22

Many facultative anaerobic bacteria undergo several physiological changes during adaptation to anaerobiosis.
- Replace oxidases with reductases for respiration
-For many organisms, latter half of the TCA cycle becomes a reductive pathway.
-For enterics and lactic acid bacteria, pyruvate dehydrogenase is replaced with pyruvate-formate lyase for oxidation of pyruvate to acetyl CoA.
-Many organisms undergo mixed acid fermentation or butanediol fermentation.

Question 23

What happens in mixed acid fermentation?

Answer 23

In mixed fermentation, bacteria utilize two or more different pathways in the terminal step of fermentation.
-Pathways used and products produced are growth condition and species-specific
-The “mixed acid” pathway makes multiple end produced and in variable amounts.

Question 24

What test do we use for mixed acid fermentation?

Answer 24

Methyl-Red test

Question 25

Who carries our Butanediol fermentation?

Answer 25

Enterobacter and Klebsiella, but can be carried out by a range of anaerobic bacteria.

Question 26

What is 2,3-butanediol?

Answer 26

A glycol that acts as a precursor to a multitude of compounds, from rubber to pesticides.

Question 27

Bacterial fermentation of 2,3-butanediol is of great interest because…?

Answer 27

Of the varied applications for 2,3-butanediol and its derivative chemical compounds which can be used for synthetic rubber, fuels, solvents, cosmetics, and food additives.

Question 28

What test do we use for butanediol fermentation?

Answer 28

Voges-Proskauer Test

Question 29

Bacteria capable of lactic acid fermentation are of significant ______ importance.

Answer 29

economic
-Production of a vast array of food, drink, and medicinal products.

Question 30

Lactic Acid fermentation:
What species are commonly used for fermentation of food products? And as what?

Answer 30

Lactobacillus species is commonly used for fermentation of food products as Lactic Acid Bacteria (LAB).

Question 31

What are Lactic Acid Bacteria (LAB)?

Answer 31

LAB are acid and salt tolerant, and each have optimal pH for growth.
They convert glucose or lactose to lactic acid.

Question 32

Metabolism by cheese-associated microbes greatly impacts…?

Answer 32

The sensory attributes of cheese:
-differences in microbial diversity result in tangible changes in flavor, odor, texture, and color.

Question 33

What are Homofermentative (or homolactic) LAB?

Answer 33

Only produce lactic acid.
-They prefer temperatures between 86-95F though they grow at lower temps as well.
-They produce flavors characterized by dairy, cream, or yogurt notes.

Question 34

Homofermentative lactate fermentation of lactic acid bacteria (LAB) such as Lactobacillus species have a primary end product of?

Answer 34

Primary end product of lactate.
Glycolysis to pyruvate = 2 ATP + 2 NADH + 2H+
These two NADH used to reduce pyruvate to 2 lactate. -Net gain per glucose = 2 lactate + 2ATP

Question 35

Heterofermentative LAB ferment glucose to produce lactic acid, but also…?

Answer 35

Acetic acid, ethanol, and carbon dioxide depending on conditions.
-They impart a sharper, more vinegar like-tang to foods. Like pickles.

Question 36

Heterofermentative lactate fermentation typically involves _________ fermentation by multiple LAB species:

Answer 36

Successional fermentation
-Leuconostoc spp
-Pediococcus spp
-Lactobacillus spp

Question 37

Heterofermentative lactate fermentation produces lactate via?

Answer 37

Via the decarboxylation and isomerase reactions of the PPP (2 NADH are produced instead of NADPH) to produce ribulose monophosphate.

Question 38

Heterofermentative lactate fermentation:
In step 5, there is something unusual. What is it?

Answer 38

An unusual reaction splits xyulose-5-P into PGALD and acetyl phosphate.

Question 39

Heterofermentative lactate fermentation:
The 3 NADH produced in the pathway are also _______ in the pathway.

Answer 39

Oxidized
-1 NADH reduced pyruvate to lactate
-1 NADH reduces acetyl phosphate to acetaldehyde
-1 NADH reduces acetaldehyde to ethanol

Question 40

The process of heterofermentative lactate fermentation of foods involves several different LAB genera and species, which dominate the culture in _______, based primarily on _______.

Answer 40

Succession ; pH

Question 41

Making pickles….
Leuconostoc mesenteroides gets to work, initiating fermentation….

Answer 41

Initiating fermentation, metabolizing sugars in the cucumber to produce lactic acid, carbon dioxide, ethanol, and acetic acid.
-This initial fermentation lowers pH from 7.0 to 4.5 the optimal pH for L. mesenteroides.
-As acids accumulate, pH continues to decrease, and more acid-tolerant bacteria take over.
-Pediococcus cerevisiae, Lactobacillus brevis, and Lactobacillus plantarum begin to proliferate outcompeting L. mesenteroides.
-These three species lower the pH further still, until the system reaches as low as pH 3.
-L. brevis and L. plantarum are left to complete the final stage of fermentation.

Question 42

What do LAB do in sourdough?

Answer 42

LAB not only act as natural leavening agents in sourdough and impart distinctive taste as result of fermentation products, but also have peptidase systems that specifically target gluten.

Question 43

Describe ethanol fermentation.

Answer 43

Glycolysis converts glucose into pyruvate, just like in respiration.
Going from pyruvate to ethanol is a two-step process.
- In the first step, a carboxyl group is removed from pyruvate and released as carbon dioxide, producing a two carbon molecule called acetaldehyde.
-In the second step, catalyzed by alcohol (ethanol) dehydrogenase, NADH passes its electrons to acetaldehyde, regenerating NAD+ and forming ethanol.

Question 44

It’s not the ethanol that kills you….

Answer 44

Acetaldehyde is a carcinogen that causes DNA and protein mutation.