Week 4: Cellular metabolism

Question 1

What are the different metabolic pathways in the human body?

Answer 1

Glycan biosynthesis and metabolism
Nucleotide metabolism
Biodegradation of Xenobiotics
Metabolism of amino acids
Metabolism of cofactors and vitamins
Biosyntehsise of secondary metabolites
Energy metabolism
Lipid metabolism

Question 2

What are the purposes of metabolic pathways?

Answer 2

Extraction of energy
Storage of fuels
Synthesis of important building blocks
Elimination of waste materials

Question 3

What is meant by homeostasis in term of cellular metabolism?

Answer 3

When concentrations of metabolites are kept at a steady state in the body.
When perturbed a new steady state must be achieved in order to maintain homeostasis.

Question 4

In what scenarios might the level of metabolites need to be altered very rapidly?

Answer 4

Need to increase the capacity of glycolysis during action
Need to reduce capacity of glycolysis after action
Need to increase capacity of gluconeogenesis after successful action

Question 5

Where do the different stages of anaerobic and aerobic respiration occur in the cell?

Answer 5

Glycolysis - cytoplasm
Anaerobic respiration - cytoplasm

Link reaction - mitochondrial matrix
Krebs/citric acid cycle - mitochondrial matrix
Oxidative phosphorylation - mitochondrial membrane

Question 6

What conditions must be met for a reaction to occur?

Answer 6

The pathway must be exergonic in the direction in which it proceeds.
The correct regulatory conditions must be present - this ensures current physiological needs are met.

Question 7

How do we prevent futile cycles in metabolism?

Answer 7

Reciprocal regulation - when the same molecule or treatment has opposite effects on the anabolic and catabolic pathway - this is very important when both reaction occur in the same compartment

This is also called a Substrate cycle - enzymes of pathways in opposite directions can control enzymes that catalyse reactions that are in the opposite direction.

Question 8

What is the process of glycolysis?

Answer 8

Glucose is catalysed by hexokinase and the hydrolysis of 2 ATP molecules to glucose-6-phosphate.
Which is converted to fructose 6-phosphate by phosphoglucose isomerase
Then converted to fructose 1,6 phosphate by phosphofructokinase 1
Is then broken down into two molecules of G3P.
Then requires the production of 2 ATP molecules and the reduction of two NAD+ to NADH to form two molecules of triose phosphate via intermediates.
Then more reactions occur including the final step which is catalysed by pyruvate kinase to produce pyruvate and 2 ATP molecules.

Question 9

What are the total inputs and outpurs of glycolysis?

Answer 9

Input: Glucose molecule, 2 ATP molecules, 2 NAD moelecules

Output: 2xpyruvate molecules, 4 ATP molecules, 2 NADH+ molecules.

Question 10

What are the four different fates of pyruvate?

Answer 10

1.Irreversibly converted to Acetyl-CoA by pryruvate dehydroganse complex - enters critic acid cycle or fatty acid synthesis

2. Irreversibly converted by oxaloacetate by pyruvate carboxylase, used as a CAC intermediate or substrate for gluconeogenesis
3. Reversibly converted to lactate (anaerobic respiration)
4. Reversibly used in ethanol production (in yeast and bacteria only).

Question 11

How is ethanol produced from pyruvate?

Answer 11

Loss of CO2 to form Acetaldehyde
Reduced by NADH (becomes NAD+) to form ethanol
is reversible

Question 12

What is the process of anaerobic respiration in humans?

Answer 12

Glycolysis occurs as normal and results in the production of 2 pyruvate molecules.
Pyruvate in reduces by NADH (becomes NAD+) to form lactate
Net ATP gain from glycolysis only - purpose is to recycled NADH to form NAD+ to be reused in glycolysis

Question 13

What is the location of the four fates of pyruvate metabolism?

Answer 13

Aerobic gluconeogenesis - liver
Aerobic cellular respiration - mitochondrial matrix
Anerobic respiration - exercising muscle or rbcs
Ethanol production - intestinal flora

Question 14

What are the different names of the citric acid cycle?

Answer 14

The krebs cycle
The tricarboxylic acid cycle

Question 15

How is the citric acid cycle utilised when glucose levels are low?

Answer 15

Muscle uses lipolysis to produce susbtances that can enter the critic acid cycle
Glucose is preserved for the brain and RBCs

Question 16

What is the link reaction?

Answer 16

Pyruvate looses a carbon dioxide and donates electrons to two molecules of NAD+ (form 2xNADH) to become acetate
Acetate combines with co-enzyme A to produce Acetyl CoA

Question 17

What happens in the citric acid cycle?

Answer 17

Acetyl-CoA (2C) enters the cycle by combining with oxaloacetate (4C) to form citrate (6C).
Undergoes a series of reactions in a cyclical pattern, involves the loss of two carbon dioxide molecules, donating electrons to three NAD+ molecules to form three NADH molecules, and donating one elctron to FADH to form FADH2, and the production of one ATP molecule by substrate level phosphorylation.
Results in the the production of oxaloacetate ready to restart the cycle

Question 18

What is the detailed list of substrates produced in the critic acid cycle?

Answer 18

1. Oxaloacetate and Acetyl-CoA combine to form citrate
2. Becomes Isocitrate
3. Loss of CO2 and electron to become a-Ketoglutarate (NADH)
4. Loss of Co2 and electron to become Succinyl CoA (NADH)
5. ATP generation (loss of phosphate) to become succinate
6. Loss of electron the produce Fumarate (FADH2)
7. Becomes Malatate
8. loss of electron to become oxaloacetate (NADH)

Question 19

What is chemiosmosis?

Answer 19

Where the movement of ions across a cell membrane creates a concentration gradient that can then be couple with ATP production - this is seen clearly with oxidative phosphorylation and H+ movement coupled with ATPsynthase.

Question 20

What is the process that occurs in oxidative phosphorylation?

Answer 20

NADH and FADH2 donate their electrons to proteins in the electron transport chain in the mitochondrial matrix
This triggers a series of redox reactions. Energy is slowly released as electrons travel down an energy gradient.
This provides the energy to actively transport the H+ ions (also released from NADH and FADH2) across the mitochondrial inner membrane from the matrix into the intermembrane space
H+ ion can then move down a concentration gradient back into the matrix through ATP synthase, this generates the energy to convert ADP and P to ATP.

Question 21

What are the different carriers /oxidoreductase enzymes in the electron transport chain is oxidative phosphorylation?

Answer 21

Complex 1: NADH dehydrogenase
Complex 2: Succinate dehydrogenase
Complex 3: Cytochrome bc1 complex
Complex 4: cytochomre c oxidase

Mobile carriers include: CoQ (ubiquiqinone)- lipid soluble carries from complex 1/2 to 3 and Cytochrome c which is water soluble carries to complex 4

Question 22

Why is FADH2 considered less efficient in energy production than NADH?

Answer 22

FADH is less efficient at donating electrons
Donates to complex 2 which does not pump protons across the membrane
Bypasses complex 1 which is used by NADH, results in less contribution to energy release hence less H+ pumped against a concentration gradient.

Question 23

What is the final electron acceptor in oxidative phosphorylation?

Answer 23

2H+ + 1/2O2 + 2e- forms water

Question 24

What is the reversal of glycolysis?

Answer 24

Gluconeogenesis - non carbohydrate substances typically converted to a pyruvate intermediate then glucose in the liver.
Used to maintain (inc) plasma glucose levels back into the healthy range.

Question 25

What enzymes ensure that glycolysis is not the exact reversal of gluconeogenesis? Contribute to recepirocal regulation

Answer 25

Glycolysis: Hexokinase convertes glucose to glucose 6 phosphate.
Phosphofructokinase 1 convertes F6P to F16biP
Pyruvate kinase converts phosphophenal pyruvate to pyruvate

Gluconeogenesis: glucose 6 phosphatase converts glucose 6 phosphate to glucose
Fructose 1,6 biphosphatase 1 converntes F16biP to F6P
Bypass reaction using pyruvate carboxylase and PEP carboxylase convert pyruvate to oxaloacetate to phosphoneal pyruvate

Hence total 3 reactions are different and regulated.

Question 26

What is the process of gluconeogenesis (from pyruvate substrate)?

Answer 26

1. In the mitochondrial matrix pyruvate carboxylase converts pyruvate to oxaloacetate
2. Converted to phosphoenolypyruvate by PEPCK enzyme.
3. Recieves an electron (from NADPH) and a P (from ATP) to produce G3P, which becomes Fructose 1,6 biphospate
4. Fructose 1,6 biphosphates enzyme converts the fructose 6 phosphate, then becomes glucose 6 phosphate
5. Finally converted by glucose 6 phosphatase to glucose

Question 27

Describe how other substances (not pyruvate) can be utilised in gluconeogenesis.

Answer 27

1. Glycerol from triglyceride breakdown - can be converted to DHAP then F16biP…. to become glucose (adipose tissue)
2. Lactate is directly oxidised (NAD+ gains e-) to become pyruvate (muscle)
3. Glucogenic amino acids can be converted to alpha ketoacids which can be converted to pyruvate (muscle)

Question 28

What is the key function of gluconeogenesis?

Answer 28

Maintain blood glucose during high fat and low carbohydrate diet to fast
Clears blood lactate from rbcs and exercising muscles

Question 29

What and how do non-glucose dervied substrates enter the citric acid cycle?

Answer 29

1) Adipose tissue - TG hydrolysed, glycerol produced can be converted to pyruvate via a DHAP intermediate then enter the links reaction

2) Muscle - protein, hydrolysed to amino acids which are then deaminated, enter at different points dependent on the number of carbon atoms 3C as pyruvate and 4C as different intermediates in citric acid cycle

A level

Question 30

What are the different factors affecting the rate of a biochemical reaction?

Answer 30

Concentration of reactants v products
Activity of the catalyst
Concentration of effectors
Temperature
pH

Question 31

What is an example of a concentration of an effector having an effect on gluconeogenesis/ glycolysis?

Answer 31

ATP concentration
High ATP concentration inhibts the committed step in glycolysis (prevents F6P being converted into F16biP)
Contrastingly increases the rate of the reverse reaction

Question 32

What can effect the activity of a catalyst?

Answer 32

Concentration of enzyme (translation v degradation)
Intrinsic activity of enzyme (depend on substrate, effector or phosphorylation state)

Question 33

What can be considered an effector in a biochemical reaction?

Answer 33

Allosteric regulators
Competing substrates
pH, ionic environment

Question 34

How does temperature increase the enyzme rate of reaction in humans?

Answer 34

Human metabolism optimal temperature is 36.8 degrees
Q10 ratio between 2-3 for most enzymes, meaning increase temp by 10 degrees increases rate of reaction by 2-3 fold
Generally, increase temp causes increased kinetic energy - increase frequency of random collision - increase successful collisions - increase rate of reaction

Question 35

How does temperature decreases the enzyme rate of reaction?

Answer 35

However above a certain temperature, hydrogen bonds (and eventually ionic bonds) break as increased vibrational energy puts pressure on bonds and the enzyme denatures, due to the active site changing shape so it is no longer complimentary to substrate, rate of reaction decreases and eventually will stop is the enzyme is completely denatured.

As more enzymes are denatured there is a greater decrease in the rate of reaction.

Question 36

What factors effect the total enzyme activity?

Answer 36

The number of enzyme molecules in teh cell
The effective activity
Modulating the activity of an existing molecule

Question 37

How do you increases the effective activity of an enzyme?

Answer 37

Extracellular signals
Transcription of specific genes
mRNA degradation
mRNA translation on ribosome
Protein degradation (ubiquitin)
Enzyme sequester in a subcellular organelle

These all effect the concentration of the active enzyme in the compartment where it is active

Question 38

How can you alter the activity of existing enzymes?

Answer 38

Enzyme binding to substrate (inc substrate conc)
Allosteric effectors
Enzyme phosphorylation (other post-translational modifications)
Enzyme combine with a regulatory protein

Question 39

Describe how the concentration of substrates effects the enzymatic rate of reaction?

Answer 39

Has a greater effect at lower concentrations of substrate

Increasing concentration - initially has a greater effect as lots of available enzymes - increases the number of enzyme-substrate complexes that can form, as increases the frequency of collisions between

Smaller effect at higher concentrations as enzyme active sites are almost saturated so increases substrate is less likely to collide with an empty active site
Has no more effect once the enzyme active sites are completely saturated - the enzyme is working at its maximum rate this is known as Vmax

Question 40

What is the importance of Km in relation to enzyme activity?

Answer 40

Also known as Michealis constant
This is the substrate concentration at which the rate of reaction if 50% of Vmax.
This is often near the physiological concentration of the substrate found naturally in the body - ensuring the enzyme is adapted to its environment as is able to work efficiently to supply metabolic demand.

Question 41

Give an overview of (de)phosphorylation of enzymes

Answer 41

Phosphorylation in catalysed by kinases
Dephosphorylation can be spontaneous or catalysed by protein phosphatases
Typically phosphorylated on hydroxyl groups of Ser/Thr/Tyr
Phosphorylation tends to increase the activity of enzymes

Question 42

How can regulatory proteins affect enzyme activity?

Answer 42

Enzyme often binds to scaffold subunit which is then also bound to be regulatory subunits
Different regulatory subunits may be able to bind with different subunits having different effects on enzyme activity

Question 43

What is the relationship in most metabolic pathways and equilibrium?

Answer 43

Most reactions are near to but not at equilibrium, and to maintain this steady state most enzymes operate at the same rate - this ensures the concentration of substances in the pathway remains relatively constant

Key enzymes operate far from equilibrium , these are considered sites of regulation and control the flow through the pathway by ensuring one metabolic pathway is favoured over another.

Question 44

How does ATP concentration act as a cellular regulator?

Answer 44

Decrease in ATP can decrease the effectiveness of ATP utilising enzymes
This is because decrease ATP leads to an increase in AMP, AMP is a potent allosteric regulator of enzymes

Question 45

How does AMP concentration effect the rate of carbohydrate and fat metabolism?

Answer 45

AMP activates AMP-activated protein kinase (AMPK)
AMPK can be activated by exercise, SNS activation, leptin and adiponectin

AMPK phosphorylates target proteins.
Shifts metabolism away from energy consuming anabolism (reduces fatty acid synthesis, glycogen synthesis, cholesterol synthesis and insulin secretion)
Shift towards energy producing processes such as gluconeogenesis in the liver, uptake of fatty acids and beta oxidation, glucose uptake, glycolysis and increased appetite (stimulates hypothalamus).

Question 46

What is meant by a regulated enzyme?
What does the activity of these enzymes tend to be like?

Answer 46

Enzymes in a metabolic pathway, which is influenced by other biomolecules, typically related to other stages in the same pathway is able to influence its own activity hence the progression through the pathway.

often ezymes in opposite pathways that have the same substrate and product (but reversed and obviously different enzymes)

Tend to operate far from equilibrium

Question 47

What enzymes are the main controllers of glycolytic flux in glycolysis?

Answer 47

Hexokinase - allows activation of glucose - largest affect on flux
Phosphofructokinase enables catabolism of activated glucose (glucose phosphate) vai glycolysis

Both these enzymes are heavily regulated and operat for from equilibrium

Question 48

What factors regulate the activity of hexokinase?

Answer 48

A nuclear protein acts as the regulatory protein, draws hexokinase IV into the nucleus when glucose concentration is low, and allows to leave the nucleus back into the cytoplasm when glucose concentration is higher.
Occurs in the liver.

Question 49

What are meant by isomers of an enzyme?
What are the features of this?

Answer 49

Isomers are different enzymes (structurally) that catalyse the same reaction
Typically share similar reaction sequences
May have different kinetic properties
Can be regulated differently

Question 50

What are the different isomers of hexokinase?
How are they different?

Answer 50

Four isomers in total
HK I is expressed in all tissues to different levels
HK IV is only expressed in the liver.

HKIV has a higher Km so is respovie to higher glucose concentration, hence is able to convert high blood glucose into glycogen stores
Is not inhibited by glucose-6-phosphate so can function at higher glucose and higher rates of glycolysis

Question 51

What is the commitment step in glycolysis?

How is it regulated?

Answer 51

Conversion of Fructose-6-phospate to frucotse 1,6 biphsophate.

ATP is a substrate and a negative regulator at high conc, do not wast glucose is plentiful ATP is available.
Inhibited by citrate
RAte increased by AMP, ADP and fructose 2,6 biphsophate (not F26biP is not a glycolytic intermediate only a regulator)

Question 52

What is the role of fructose 2,6 diphosphate as a regulator?

Answer 52

Is not a glycolytic intermediate
Increase phosphofructokinase (glycolysis)
Inhibit fructose 1,6 biphosphatase (gluconeogenesis)

Question 53

What conditions favour gluconeogenesismetabolic pathway?

Answer 53

Low AMP
High ATP/citrate