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Cellular and Molecular Biology > Lecture 29: Metabolic Pathways and Glycolysis > Flashcards

Lecture 29: Metabolic Pathways and Glycolysis Flashcards

25/11/2024

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1
Q

What were the beliefs surrounding respiration in the late 1800s

A
  • Respiration essential for life and requires living cells (vitalism)
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2
Q

Maria Mikhàilovna Manàsseina in the 1890s

A
  • Manàsseina observed that fermentation was an enzymatic process that didn’t require live cells to be present at all/vitalism
  • Buchner brothers later observed fermentation in cell-free extracts (crushed yeast)
  • Did so by trying to preserve active yeast extract using sucrose
  • These experiments led to the end of vitalism
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3
Q

Did Eduard Buchner fail to credit Manasseina?

A

Yes, leading him to getting the nobel prize in 1907

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4
Q

By 1900..

A

.. it was shown that muscle extract is also active but during fermentation produces lactate, and not alcohol.

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5
Q

By 1940..

A

–the entire glycolysis pathway had been pieced together through the work of many biochemists. Particularly Harden Young, who discovered some of the key components of the glycolytic pathway

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6
Q

Why is glycolysis sometimes referred to as the Embden-Meyerhof pathway?

A

Because both Embden and Meyerhof played a key role in piecing together the components of glycolysis.

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7
Q

Describe Harden and Young’s Dialysis of Yeast Extract experiment for discovering the process of glycolysis

A
  • Harden and Young were investigating how yeast extracts could ferment glucose into alcohol and carbon dioxide. Earlier, Eduard Buchner had discovered that yeast extracts (cell-free) could perform fermentation, proving that fermentation did not require living cells, but rather the action of enzymes. Harden and Young extended this work to investigate the chemical details of the process.
  • Harden and Young put yeast lysate in a bag and put it in a buffer for several hours
  • They found that bigger molecules stayed in the bag, but smaller molecules diffused through the membrane. This separated the yeast into 2 extracts.
  • They called the large molecules zymase and the small molecules co-zymase.
  • They discovered that neither fraction alone could ferment glucose into ethanol and carbon dioxide. However, when the two fractions were recombined, fermentation resumed, producing carbon dioxide and ethanol.
  • Hence, they concluded that both the small molecules (from the dialyzable fraction) and the enzymes (from the non-dialyzable fraction) were essential for fermentation. And that fermentation involves cofactors, which are small molecules that work alongside enzymes to drive the reaction.
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8
Q

What is zymase?

A

large molecules (non-dialysable)
inactivated by heat (proteins, enzymes)

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9
Q

What is cozymase?

A

small molecules (dialysable)
heat stable (substrates and coenzymes)

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10
Q

What are enzymatic cofactors and coenzymes?

A

Non protein substances that are required for the catalytic activity of some enzymes

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11
Q

What 2 things can cofactors and coenzymes be?

A
  • Inorganic
  • Metal ions such as: Mg2+, Zn2+, Mn2+, etc.
  • Organic. E.g. Nicotinamide adenine dinucleotide (NAD+) and Flavin adenine dinucleotide (FAD)
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12
Q

What are NAD+ and FAD both derived from?

A

Both derived from B vitamins

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13
Q

Is it true that enzymatic cofactors and coenzymes are small and heat stable?

A

Yes

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14
Q

Harden and Young: Phosphate is limiting glycolysis

(Their second experiment)

A
  • Looked at how much carbon dioxide was being evolved over time during glucose fermentation and using yeast extract.
  • Found that increasing the sucrose concentration (sucrose would consequently be hydrolysed into glucose) had no effect on increasing the carbon dioxide concentration.
  • This is because despite sucrose being the substrate that is used up, sucrose is not the limiting factor.
  • They then established that inorganic phosphate is actually the limiting factor in glycolysis.
  • It was later shown that NAD+ was also used up in glycolysis.
  • This was an in vitro approach
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15
Q

What is the glycolysis equation?

A

Glucose + 2 ADP+ 2 Pi+ 2 NAD+ -> 2 pyruvate + 2ATP +2NADH+2H++2H2O

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16
Q

What does the in vitro approach include?

A

Taking cells, adding buffer, and homogenising them

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17
Q

What are the advantages of using the in vitro approach to study metabolic pathways?

A
  • Allows for the study of a purified enzyme in isolation
  • Allows for completely defined (and readily modifiable) conditions
  • Allows for direct, quantitative results
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18
Q

What are the drawbacks of using the in vitro approach to study metabolic pathways?

A
  • Results in a loss of compartmentation, and a loss of spatial and temporal organisation
  • Can result in the instability or degradation of key components e.g. lack of ATP consumption in H&Y extracts prevented Pi release, limiting glycolysis
  • We can’t know for sure that the properties of a cell-free extract accurately reflects the properties of intact cells
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19
Q

Describe the in vivo approach

A

Assays and indications
e.g., isotopes: D2O reveals hydrolysis, radioactive isotopes can trace reactions.

Can involve modifying systems by adding inhibitors or mutations

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20
Q

What are the advantages of the in vivo approach?

A
  • Looking at cells and organisms gives a better reflection of the whole system
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21
Q

What are the disadvantages of the in vivo approach?

A
  • Involves careful interpretation due to there being many variables
  • Can be hard to get quantitative data
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22
Q

Why do we focus on glucose metabolism?

A
  • Because it’s central to our energy metabolism and creates the building blocks for other molecules.
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23
Q

What is metabolism organised in?

A

Pathways

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24
Q

What are the 3 stages of metabolism?

A
  • Stage I
  • Stage II
  • Stage III
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25
Q

What does Stage I of energy metabolism involve?

A
  • Converting macromolecules, such as proteins, into smaller building blocks, such as amino acids.

-There is no useful energy production in this stage

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26
Q

What does Stage II of energy metabolism involve?

A
  • The conversion of many products of Stage I into Acetyl-CoA .
  • A small amount of ATP is produced
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27
Q

What does Stage III of energy metabolism involve?

A
  • The oxidation of acetyl CoA

-

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28
Q

What is catbolism?

A

involves the breakdown of complex molecules into simpler ones, releasing energy in the process

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29
Q

What is anabolism?

A

Involves the involves the synthesis of complex molecules from simpler ones, requiring an input of energy.

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30
Q

Why do we need multi-step metabolic pathways?

A
  • Allows for the release of energy to be more controlled. This allows for energy to be conserved, as energy can be harvested little by little in a much more controlled way.

-

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31
Q

What are the 6 key types of reactions?

A

Redox reactions

Group transfer

Hydrolysis

Addition or removal of functional groups

Isomerisation

Ligations

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32
Q

Consider metabolic pathways with the reaction types in mind

A

Consider metabolic pathways with the reaction types in mind

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33
Q

Do many metabolic pathways crossover? Give an example of this

A

Yes

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34
Q

Is it true that normally the key steps of metabolic pathways are: irreversible, have a very strongly negative ΔG, and so are used as control points?

A

Yes

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35
Q

Why can’t anabolic reactions simply be be the reverse of catabolic reactions?

A

Because normally the key steps are the irreversible ones (i.e. strongly negative ΔG) –control points

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36
Q

What do most metabolic pathways usually include?

A

A small number of intermediates

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37
Q

Is it true that a small set of certain cofactors carry out a wide range of metabolic tasks?

A

Yes

38
Q

What are the key cofactors in energy metabolism?

A

ATP

NAD+, FAD

NADPH

CoA

39
Q

Name a cofactor that is an energy carrier

A

ATP

40
Q

Name cofactors that are electron carriers and used for the oxidation of fuels

A

NAD+, FAD

41
Q

Name cofactors that are electron carriers and used for reductive biosynthesis

A

NADPH

42
Q

Name a cofactor that is a 2-carbon groups carrier

A

CoA (coenzyme A)

43
Q

Is it true that cofactors are all stable molecules?

A

Yes, and they react slowly in the absence of enzymes

44
Q

What does NAD stand for?

A

Nicotineamide Adenine Dinucleotide

45
Q

What does FAD stand for?

A

Flavin Adenine Dinucleotides

46
Q

What is NAD derived from?

A

Derived from Niacin/vitamin B3

47
Q

Draw the structure of NAD

A
48
Q

What is FAD derived from?

A

Derived from Riboflavin/vitamin B2

49
Q

Draw out FAD

A
50
Q

Do NAD and FAD have very similar structures?

A

Yes

51
Q

Draw out the structure of NAD

A
52
Q

Draw out the structure of FAD

A
53
Q

What is the only difference between NAD and NAPD?

A

NAPD has a phosphate group, ensures that they are recognised by different enzymes

54
Q

What are key points to ask of metabolic pathways whenever you come across them?

A

Where in the cell does it occur?

Is it linear or branched?

What are the entry points?

What are the end products?

Which other pathways is it linked to?

How does it work step-by-step?

What are the irreversible steps, i.e. the key control points?

How is it regulated?

54
Q

What is the respiration equation?

A

C6H12O6 + 6O2 → 6CO2 + 6H2O

55
Q

What is the first step of respiration?

A

Glycolysys

56
Q

In summary, what is glycolysis?

A

A series of reactions starting with glucose and ending with pyruvate resulting in a small gain of ATP (2 molecules ATP for each glucose molecule)

57
Q

What is glucose oxidised to in glycolysis?

A

2 X Pyruvate

58
Q

Is glycolysis aerobic or anaerobic?

A

Anaerobic

59
Q

Why is glycolysis anaerobic?

A

Because the process evolved before the atmosphere contained significant O2

60
Q

How is ATP produced in glycolysis?

A

Free energy released by oxidation allows ATP production from ADP + Pi

61
Q

Is the pathway of glycolysis linear or branched?

A

Linear

62
Q

Where is glycolysis located?

A

In the cytosol

63
Q

What happens to the intermediates in glycolysis?

A

They are phosphorylated- the first phosphorylation makes sure that glucose stays in the cell.

64
Q

How many steps are there in glycolysis?

A

10 steps each catalysed by a different enzyme

65
Q

What is the net gain of ATP in glycolysis?

A

2 per glucose.

  1. Investment Stage (Energy Input)
    2 ATP are used: You spend 2 ATP to prepare glucose for breaking down.
  2. Payout Stage (Energy Generation)
    Oxidation: Some of the molecules are oxidized (lose electrons), which helps produce energy.
    4 ATP are made: You get 4 ATP from transferring phosphate groups to ADP (through substrate-level phosphorylation).
    2 NADH are made: NAD+ picks up electrons and becomes NADH during oxidation.

Net Gain:
2 ATP: After spending 2 ATP and making 4 ATP, you end up with a net gain of 2 ATP.
2 NADH and 2 pyruvate are also produced.

66
Q

How many regulatory steps are there in glycolysis? Label them

A

3

67
Q

Step 1 regulatory step

A
  • Hexokinase, feedback inhibition
  • Prevents glucose from escaping from cell
  • However, not main regulatory step because there are other entry points of glucose, and glucose is needed in other pathways.
68
Q

Step 3 regulatory step

A
  • Key regulatory step for glycolysis
  • PFK enzyme, responds to enegry leveld (of AMP and ATP) and makes sure that there is sufficient metabolic intermediates
69
Q

Step 10 regulatory step

A
  • Feedforward activation
  • Not the main regulatory step, but needed as a metabolic branch point, important in other metabolic pathways.
70
Q

High levels of glucose-6-phosphate inhibit hexokinase through a feedback inhibition mechanism. Is this statement true???

A

Yes

71
Q

What is needed to generate ATP for muscles to use?

A

Glycolysis

72
Q

What happens when there’s a high ATP concentration in the muscles?

A
  • Enzyme PFK inactivated
  • G-6-P accumulate
    If not used elsewhere
  • Enzyme HK inactivated
73
Q

What happens when there’s a high AMP concentration in the muscles?

A
  • PFK activated
  • G-6-P used up
  • HK activated
74
Q

Why is the control of metabolic pathways more complicated in liver than in muscles an?

A

Because the liver has more functions than the muscles. For example, Glycolysis needed to generate intermediates for biosynthesis and (energy)
Liver must regulate blood glucose levels

75
Q

Are ATP and AMP regulated in liver as well, but levels don’t vary that much?

A

Yes

76
Q

Apart from ATP and AMP what 2 other substances are also regulated in the liver?

A

Citric acid and Fructose-2,6-P

77
Q

What is citric acid an intermediate in?

A

The citric acid cycle

78
Q

Fructose-2,6-P increases when blood glucose is high
Technically responds to high F-6-P in cell (feed forward)
Reduces inhibition by ATP

A

Fructose-2,6-P increases when blood glucose is high
Technically responds to high F-6-P in cell (feed forward)
Reduces inhibition by ATP

79
Q

Apart from glucose, what other dieatry sugars can be used in glycolysis?

A

Galactose and Fructose

80
Q

Show how galactose can be metabolised at glucose in glycolysis through drawing a diagram

A
81
Q

What is our source of galactose in our diet?

A

Lactose (galactose comes from lactose)

82
Q

Why are people lactose intolerant?

A

Because they lack the enzyme that
breaks down lactose

83
Q

Why do people have Galactosemia?

A

Because they cannot break down galactose (rare)

84
Q

How is Galactosemia treated?

A

Treated by avoidance of galactose in diet

85
Q

Draw a diagram and show the process of how Fructose is converted into Glyceraldehyde3-P

A
86
Q

What stage of glycolysis does fructose enter?

A

Stage 2 as Glyceraldehyde3-P. i.e. after the main regulatory step catalysed by PFK

87
Q

What sugar can the naked mole rat survive on?

A

Fructose

88
Q

What enzyme is glycolysis mainly regulated by?

A

PFK

89
Q
A

Cellular and Molecular Biology (33 decks)

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