Metabolism 2 + 3

Question 1

What happens to part of the energy released during oxidation reactions?

Answer 1

-coupled to generation of activated carrier molecules
-molecules then drive endergonic reactions

Question 2

At equilibrium, why is there a higher concentration of ADP and Pi?

Answer 2

-negative charges on phosphates repel
-entropy increases
-water stabilises the product - more interactions with ADP + Pi
-free Pi stabilised by resonance structures not possible when bound to ATP

Question 3

How do you calculate ΔG?

Answer 3

ΔG = RT ln(mass action ratio/K)

Question 4

What is the mass action ratio?

Answer 4

The actual concentration ratio of products to reactants under a particular condition

Question 5

For a dead cell, what is the ΔG for when the mass action ratio is less than, equal to or greater than K?

Answer 5

-< K, ΔG = -ive
-=K, ΔG = 0
->K, ΔG = +ive

Question 6

In a living cell, is the mass action ratio less than, equal to or greater than K?

Answer 6

< K , ΔG = -ive

Question 7

What defines the capacity of a reaction to do work?

Answer 7

The extent to which the actual concentration ratio of products to reactants (mass action ratio) is displaced from equilibrium

Question 8

What are features of cofactors?

Answer 8

-kinetically stable - enzymes can control flow of free energy + reducing power
-thermodynamically stable
-all react slowly with O2/H2O in absence of enzyme catalyst

Question 9

What is coupled in anabolic pathways and some steps in catabolism?

Answer 9

-endergonic reactions (+ΔG) to exergonic reactions (-ΔG)
-net -ΔG

Question 10

Where does glycolysis take place?

Answer 10

Cytoplasm

Question 11

What is the net gain of ATP and NADH in glycolysis?

Answer 11

Net gain of 2 ATP + 2 NADH

Question 12

What are the steps of glycolysis?

Answer 12

1. glucose phosphorylation
2. isomerisation to fructose
3. second phosphorylation
4. cleavage
5. conversion of DHAP
6. oxidation of GAP
7. first phosphate transfer to ADP
8. isomerisation to 2-phosphoglycerate
9. removal of water
10. second phosphate transfer

Question 13

Which steps in glycolysis are repeated?

Answer 13

-steps 6-10 occur twice
-each glucose molecule produces 2 pyruvate

Question 14

Describe the first step of glycolysis

Answer 14

-2 ATP hydrolysed to release energy
-terminal phosphate transferred from ATP to 6th carbon on glucose to form G6P using enzyme hexokinase
–ive charge on Pi group means G6P trapped in cell
-keeps glucose conc. in cell low so increased glucose uptake

Question 15

Describe hexokinase

Answer 15

-glucose binding causes conformational change
-favours direct transfer of Pi from ATP to glucose + prevents ATP hydrolysis
-active site contains aspartate - deprotonates C6 hydroxyl group on glucose
-deprotonated O- acts as nucleophile + attacks gamma Pi group of ATP
-Pi transferred directly to glucose without hydrolysis by water

Question 16

Which reactions are coupled in glycolysis?

Answer 16

-ATP hydrolysis with addition of Pi to glucose
-results in net -ΔG
equilibrium ratio of [Glu]/[G6P] changed by factor of 10^5

Question 17

Describe the second step of glycolysis

Answer 17

-G6P isomerised by phosphoglucose isomerase to fructose-6-phosphate (F6P)
-forms a ketose sugar from an aldose sugar - needed for step 4
-reaction readily reversible under cellular conditions

Question 18

Describe the third step of glycolysis

Answer 18

-F6P phosphorylated by phosphofructokinase to fructose-1,6-bisphosphate (F1,6P) using ATP
-Pi group further destabilises sugar - promotes cleavage in step 4
-entry of sugars into glycolysis controlled via allosteric regulation of phosphofructokinase

Question 19

Describe step 4 of glycolysis

Answer 19

-F1,6P is cleaved by aldolase, producing 2 3C sugars, DHAP + GAP
-DHAP converted to GAP as only GAP proceeds through glycolysis

Question 20

Why is the isomerisation to fructose in step 2 important?

Answer 20

-ensures a 3:3 split rather than a 2:4 split
-that would require 2 separate pathways

Question 21

Describe step 5 of glycolysis

Answer 21

-DHAP converted to a second GAP molecule by triose phosphate isomerase (TIM)
-TIM is a kinetically perfect enzyme
-TIM suppresses formation of toxic intermediate methyl glyoxal from the enediol intermediate
-does this by movement of 10 AA loop region over active site that blocks exit of enediol until GAP formed

Question 22

Describe step 6 of glycolysis

Answer 22

-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) uses NAD+ to oxidise GAP to form NADH
-enzyme couples oxidation to transfer of Pi group to the sugar to form 1,3BPG (has high phosphoryl transfer potential)
-the first energy generating step of glycolysis

Question 23

How is NADH made and what is it used to make?

Answer 23

-energy released from carbon oxidation used to form NADH via transfer of a H- to NAD+
-used by electron transfer chain in mitochondria to make ATP by oxidative phosphorylation

Question 24

Describe step 7 of glycolysis

Answer 24

-Pi group transferred from 1,3BPG to ADP by phosphoglycerate kinase - forms ATP + 3-phosphoglycerate (3PG)
-substrate level phosphorylation - Pi transferred from phosphosugar e.g 13BPG to ADP
-second energy generating step
-standard ΔG is -ive
-Mg2+ in active site activates ADP for reaction

Question 25

Describe step 8 of glycolysis

Answer 25

-remaining phosphoester linkage transferred by phosphoglycerate mutase from C3 to C2
-forms 2-phosphoglycerate
-facilitates removal of water in step 9

Question 26

Describe step 9 of glycolysis

Answer 26

-water removed from 2-phosphoglycerate by enolase, creating a molecule of phosphoenolpyruvate
-2 phosphoglycerate + PEP contain same amount of metabolic energy
-enolase reaction rearranges substrate into a form from which more of this potential energy can be released upon phosphoryl transfer

Question 27

Describe step 10 of glycolysis

Answer 27

-transfer of phosphate group from step 9 by pyruvate kinase to ADP forms ATP + pyruvate
-the 3rd energy generating step + another substrate level phosphorylation
-reaction runs far from equilibrium + essentially irreversible
-pyruvate conc. kept low by product removal

Question 28

When does fermentation occur?

Answer 28

-in the absence of O2
-further pyruvate oxidation can’t occur

Question 29

What accumulates when O2 is absent, why and why is this bad?

Answer 29

-NADH
-inhibition of electron transport
-NAD+ shortage caused + glycolysis inhibited

Question 30

How is the NAD+ shortage prevented when O2 is absent?

Answer 30

-NADH used to reduce pyruvate to lactate or ethanol
-NAD+ regenerated to restore redox balance
-ATP production continues