Lecture 8 - principles of bacterial aerobic cellular respiration

Question 1

Growing cells

Answer 1

new biomass dominates total energy requirements

Question 2

Non growing cells

Answer 2

repair and replacement is all that is possible

Question 3

Proton motive force

Answer 3

gradient of both concentration and charge that is required even during non growth

Question 4

3 key molecules that participate and determine the majority of catabolic and anabolic reactions

Answer 4

ATP, NADH and NADPH

Question 5

Active vs passive transport

Answer 5

Active transport accumulates substrates rapidly and against concentration gradient, but has a higher energy cost than passive

Question 6

3 transporters are

Answer 6

ABC family, MFS family, group translocation

Question 7

Protein synthesis has the highest…

Answer 7

Protein synthesis has the highest energy requirement, making new amino acids monomers however would require more energy than protein synthesis

Question 8

How do food sources get converted into NADH?

Answer 8

Amino acids, glucose, glycerol and fatty acid are monomers

Variety of different nutrient sources that cell can take up such as proteins, sugars and lipids. Idea behind cell metabolism in general is that we are trying to conserve backbones so that we can used conserved pathways to operate on because then you do not have to make millions of different proteins that are really hard to make so you want to make as few proteins as possible whilst still maintaining flexibility

Monomers are entered into various parts of glycolysis or the citric acid cycle//breakdown products ultimately enter the CAC and can be entered into different parts of this conserved pathway depending on the molecule or pathway used initially (see diagram)

Specialised pathways isolate the conserved carbon backbone from molecules to enter it into either glycolysis or TCA cycle

Question 9

Monomers and how they are converted to NADH

Answer 9

Monomers are entered into various parts of glycolysis or the citric acid cycle//breakdown products ultimately enter the CAC and can be entered into different parts of this conserved pathway depending on the molecule or pathway used initially (see diagram)

Question 10

Sugars/monosaccharides conversion

Answer 10

(6 carbon rings only)

Converted to glucose-6-phosphate or early precursors of glycolysis

Question 11

Amino acids conversion

Answer 11

Amino (NH2) group must be removed. Variable entry into TCA

Primarily transaminate NH2 -> glutamate - key in CAC, example of how a specific amino acid can be entered into the CAC which means all the stuff that happened in glycolysis is missed and doesn’t happen and the alternative pathways need to be used

Glutamate alpha-ketoglutarate is tightly regulated by both TCA cycle and urea cycle. Deamination removes excess NH2

Question 12

Fatty acids conversion

Answer 12

Glycerol can enter into glycolysis with only two enzyme reactions

Beta oxidation: flexibility removes acetyl-CoA or propionyl-CoA from any length fatty acids. Both enter into TCA

Question 13

Two phases of glycolysis

Answer 13

Energy investment phase and the energy payoff phase

Question 14

Glycolysis define

Answer 14

Glycolysis is the first of the main metabolic pathways of cellular respiration to produce energy in the form of ATP. … Overall, the process of glycolysis produces a net gain of two pyruvate molecules, two ATP molecules, and two NADH molecules for the cell to use for energy

Question 15

Energy investment phase

Answer 15

Put in energy

ATP is used to phosphorylate sugar : unstable intermediate

6C sugar ring is broken into two 3C chains

Use ATP to further phosphorylate G6P and modify it into unstable intermediates, essentially glucose is split into two smaller things which go through the rest of the pathway twice

Question 16

Energy pay off phase

Answer 16

Oxidations and (eventual) removal of phosphate produced ATP and NADH

Subsequent oxidation and reduction reactions to essentially create ATP or NADH, can create ATP directly or can create NADH which is involved in some catabolic steps and then will be used by respiration later

Question 17

For each molecule of glucose…

Answer 17

2 pyruvate are produced

Question 18

What happens to the 2 pyruvate produced during glycolysis?

Answer 18

these are then entered into the CAC and essentially the idea is that it is going around in a forward (oxidative) direction and then adding in acetyl-CoA (can come from beta oxidation but pyruvate usually gets converted into acetyl-CoA) and this gets added into some of the conserved intermediates, essentially adding carbons on to the 4C molecule and then later removing carbons to generate NADH molecules

Full cycle operates twice per glucose in glycolysis

Many reactions are reversible

Aerobically the favourable direction is oxidative (forward running cycle)

Driven by dehydrogenases - enzymes that perform oxidation reactions that can produce lots of these energy currencies

Question 19

NET products produced during glycolysis

Answer 19

2 pyruvate, 2 ATP, 2 NADH

Question 20

ATP formed and used in glycolysis

Answer 20

4 ATP formed, 2 ATP used, therefore net gain of 2 ATP

Question 21

Products produced from pyruvate dehydrogenase (PD)

Answer 21

1 NADH

Question 22

Products produced during 1 full cycle of TAC

Answer 22

3 NADH
1 GTP (can be converted into 1 ATP)
1 FADH2

Question 23

Total products from the entire process ( 1x glycolysis, 2 x PD, 2 x TCA)

Answer 23

10 NADH, 2 FADH2, 2 ATP and 2 GTP

Question 24

The two major routes of ATP production

Answer 24

Substrate level phosphorylation and oxidative phosphorylation

Question 25

Substrate level phosphorylation

Answer 25

Making ATP as a consequence of cytoplasmic reactions Glycolysis and TCA possess such reactions Low efficiency but simpler and not limited by electron acceptor Eventually limited without regenerating NAD+ (e.g. fermentation) - linked to a lot of cytoplasmic reactions that need NAD+ Essentially it is any process in the cytoplasm of the cell that produces ATP, get very low amounts of ATP but can make smaller amounts of proteins in this process

Question 26

Oxidative phosphorylation

Answer 26

Oxidative phosphorylation is the process by which ATP is synthesized as the result of electron transport driven by the oxidation of a chemical energy source. Using reducing power (combining NADH and FADH2) to generate ATP Electron transport chain and ATP synthase For ETC - oxidises our reducing power, takes the electrons off things like NADH and by taking these electrons down energy levels we make a proton pump, essentially the electron is put on oxygen and this is why we breathe oxygen as humans in order to power this process and get a proton gradient which is then used to power another enzyme called ATP synthase (enzyme that produces ATP) High efficiency but complex and limited by electron acceptor Very efficient system and is goof for organisms that require a lot of ATP or for organisms that need to use their carbon source efficiently so indeed persisters and cells that grow slow are found to rely on these processes therefore understand the importance of PMF in stressful environments Regenerates NAD+ as part of its activity - allows catabolic reactions to continue which is a good way of dealing with the build up of too much reducing power Linked by proton motive force Also known as cellular respiration

Question 27

OXPHOS produces up to _____ times _____ ATP than SLP

Answer 27

10 times more

Question 28

ATP is required for

Answer 28

anabolic processes (polymer synthesis)

Question 29

Growth rate of bacteria depends on

Answer 29

ATP yields because you need ATP for anabolic processes

Question 30

Bacterial and archaeal ETCs vs mitochondrial ETC

Answer 30

Although some bacterial and archaeal ETCs resemble the mitochondrial chain, they are frequently very different. First, bacterial and archaeal ETCs are located primarily within the plasma membrane. Furthermore, some Gram-negative bacteria have ETC carriers in the periplasmic space and even the outer membrane. Bacterial and archaeal ETCs also can be composed of different electron carriers, employ different terminal oxidases, and be branched. That is, electrons may enter the chain at several points and leave through several terminal oxidases. Bacterial and archaeal ETCs also may be shorter, resulting in the release of less energy (and the transport of fewer protons across the membrane). Although microbial ETCs differ in details of construction, they operate using the same fundamental principles.

Question 31

Order used of reactions

Answer 31

from the first one used ... aerobic respiration (oxygen at high pressure), micro aerobic respiration (oxygen at low oxygen pressure), anaerobic respiration (nitrate), anaerobic respiration (fumarate) and fermentation (acetyl-CoA)

Question 32

Aerobic respiration - Oxygen ( high pO2) - MAX YIELD

Answer 32

38

Question 33

Microaerobic respiration - oxygen (low pO2) - MAX YIELD

Answer 33

15

Question 34

Anaerobic respiration - Nitrate - MAX YIELD

Answer 34

15

Question 35

Anaerobic respiration - Fumarate - MAX YIELD

Answer 35

12

Question 36

Fermentation - acetyl-CoA - MAX YIELD

Answer 36

3

Question 37

Reactions that are oxidative phosphorylation and substrate level phosphorylation

Answer 37

aerobic respiration (oxygen at high pressure), micro aerobic respiration (oxygen at low oxygen pressure), anaerobic respiration (nitrate), anaerobic respiration (fumarate)

Question 38

Reactions that are only substrate level phosphorylation

Answer 38

Fermentation - acetyl-CoA

Question 39

Why is oxygen so important in OXPHOS?

Answer 39

Redox (reduction) potentials Molecules can inherently have high energy electrons within them Main principle behind OXPHOS Electrons in redox active molecules have an inherent energy, measured as a voltage Tendency of a molecule to accept or release electrons More negative = more likely to release electron. Electron in higher energy orbital/state (itself will be oxidised and it will reduce something else) Energy release from transferring electrons can be used to do work, coupled to proton pumping Oxygen is the lowest energy electron acceptor in biological systems

Question 40

Oxygen is the ______ _______ electron acceptor in biological systems

Answer 40

lowest energy

Question 41

Redox potentials

Answer 41

More negative = more likely to release electron. Electron in higher energy orbital/state (itself will be oxidised and it will reduce something else)

Question 42

Aerobic respiration

Answer 42

oxygen is used as terminal electron acceptor

Question 43

Anaerobic respiration

Answer 43

no oxygen available so bacteria use an alternative electron acceptor

Question 44

Electron transport chain

Answer 44

Series of membrane bound proteins AND cofactors Catalyse the redox reactions that usually make a PMF Bacteria = highly modular, but the mitochondrial configuration is the most efficient. Flexible

Question 45

Key features of ETCs

Answer 45

1 - A membrane = a physical barrier is needed to create a concentration gradient 2- Oxidative protein complexes that liberate electrons from reducing power and may pump H+ 3 - Cofactors that transfer electrons between enzymes 4 - Reductive protein complexes that finally transfer electrons to a terminal electron acceptor and may pump H+

Question 46

Electron liberating complexes list

Answer 46

``` Complex I Complex II Quinone Cytochrome C Complex III Complex IV ```

Question 47

Complex I

Answer 47

Complex 1 - NADH dehydrogenase. Large multi-subunit enzyme Oxidises NADH and reduced coenzyme q (ubiquinone) Induces structural/conformational changes that pump proton s Electron liberating complexes

Question 48

complex II

Answer 48

Complex II - Succinate dehydrogenase (none protein pumping enzyme), the same enzyme from CAC FAD is in fact covalently bound intermediate. Succinate oxidised and quoin directly reduced Generally does not pump protons Electron liberating complexes

Question 49

Quinone

Answer 49

Small lipophilic chemical with aromatic headgroup that accepts electrons Small molecule that freely diffuses through the membrane at least by the standard model to fo to the terminal acceptors on the enzyme Many types with different redox potentials e.g. coenzyme Q / ubiquinone E.g. vitamin K2/ Menaquinone Called a quinol when reduced Electron transferring cofactors

Question 50

Cytochrome C

Answer 50

Cytochrome = protein with heme Small protein that contains a heme group that accepts electrons Generally only associated with oxidase enzymes that perform oxygen reduction Not to be confused with other heme containing enzymes like cytochrome P450, that perform reactions Electron transferring cofactors

Question 51

Complex III

Answer 51

Cytochrome bcI. Transfer electrons between quinol and cytochrome C Indirectly transfer protons using electron carrier acid/base chemistry (not a proton pump) Linked to complex IV, in many bacteria they are a super complex Electron donating complexes

Question 52

Complex IV

Answer 52

Cytochrome C oxidase. Electrons from cytochrome C used to terminally reduce oxygen Protons taken up during catalytic intermediate and pumped Electron donating complexes

Question 53

ETC features - a membrane

Answer 53

A membrane = a physical barrier is needed to create a concentration gradient

Question 54

ETC features - oxidative protein complexes

Answer 54

Oxidative protein complexes that liberate electrons from reducing power and may pump H+

Question 55

ETC features - cofactors

Answer 55

Cofactors that transfer electrons between enzymes

Question 56

ETC features - reductive protein complexes

Answer 56

Reductive protein complexes that finally transfer electrons to a terminal electron acceptor and may pump H+