Enzymes of citric acid cycle

Question 1

What enzyme is involved in the conversion of acetyl-CoA and oxaloacetate to citrate

Answer 1

Citrate synthase

Question 2

Describe the action of citrate synthase

Answer 2

1. Has open and closed forms
2. Oxaloacetate binds which generates a conformational change producing the acetyl-CoA binding site
3. It also seals off the solvent’s access to the bound oxaloacetate
4. Induced fit model of substrate binding- similar to hexokinase

Question 3

What is the enzyme involved in the conversion of citrate to Isocitrate

Answer 3

Aconitase

Question 4

Describe the action of aconitase

Answer 4

1. Aconitase can distinguish between optical isomers of citrate
2. It contains a [4Fe-4S] iron-sulfur cluster which is required for catalytic activity- associated with redox processes
3. Fe atom is thought to coordinate the OH group to facilitate its elimination
4. Aconitase catalyses the stereospecific trans addition of OH- and H+ across the double bond to form specific optical isomer

Question 5

What inhibits aconitase

Answer 5

1. Fluorocitrate

2. Inhibits transport of citrate across mitochondrial membrane as well

Question 6

What is the enzyme involved in the conversion of isocitrate to 2-oxoglutarate

Answer 6

1. Isocitrate Dehydrogenase

Question 7

Describe the action of isocitrate dehydrogenase

Answer 7

1. Mammalian tissues contain two different forms
2. Requires Mn2+ or Mg2+ cofactor and NAD+
3. Oxidises isocitrate to a ketone (oxalosuccinate)
4. Then decarboxylation to succinyl-CoA

Question 8

What is the enzyme involved in the conversion of 2-oxoglutarate to succinyl-CoA

Answer 8

1. 2-oxoglutarate dehydrogenase complex

Question 9

Describe the action of 2-oxoglutarate dehydrogenase complex

Answer 9

1. Involves multi-enzyme complex with 3 enzymes
2. E1- 2-oxoglutarate decarboxylase
3. E2- lipoyl-succinyl transferase
4. E3- dihydrolipodamide dehydrogenase
5. Similar mechanism as pyruvate dehydrogenase complex

Question 10

What is the enzyme involved in the conversion of succinyl-CoA to succinate

Answer 10

1. succinate thiokinase

Question 11

Describe the action of succinate thiokinase

Answer 11

1. Hydrolyses high energy compound with coupled synthesis of high energy NTP
2. GTP- produced in mammals, ATP in plant and bacteria
3. Succinyl-CoA reacts with Pi to form succinyl phosphate and CoA
4. Succinyl phosphate’s phosphoryl group is transferred to an enzyme His residue- releasing succinate
5. THe phosphoryl group on the enzyme is transferred to GDP forming GTP

Question 12

What is the enzyme involved in the conversion of succinate to fumarate

Answer 12

1. Succinate dehydrogenase

Question 13

Describe the action of succinate dehydrogenase

Answer 13

1. Contains FAD- electron acceptor
2. FAD is covalently bound to an enzyme His residue
3. Covalent link is unusual
4. It is reoxidised from FADH2 by the electron transport chain
5. Why succinate dehydrogenase is only membrane bound enzyme

Question 14

What is the enzyme involved in the conversion of fumarate to malate

Answer 14

1. Fumarase

Question 15

What is the enzyme involved in the conversion of malate to oxaloacetate

Answer 15

1. Malate dehydrogenase

Question 16

Describe the action of malate dehydrogenase

Answer 16

1. Malate dehydrogenase has a +delta G
2. Concentration of oxaloacetate is low
3. But first step in cycle is very exergonic
4. Cycle allows citrate formation to be exergonic even at low concentrations of oxaloacetate