Biochem - the citric acid cycle

What does the citric acid provide for the cell?

• Source of precursor molecules

• Storage forms of fuels

• Building blocks of amino acids

• Porphyrins - cholesterol/haem

Where do glycolysis and the citric acid cycle take place in the cell?

• Glycolysis takes place in the cytoplasm of the cell

• The citric acid cycle takes place in the mitochondrial matrix of the cell

• Pyruvate from glycolysis has to cross both mitochondrial membranes through a specific carrier protein

What happens during the ‘Link reaction’?

• Form acetyl CoA from pyruvate

• Pyruvate is transported into the mitochondrial matrix

• Oxidatively decarboxylated to acetyl CoA (loses CO2)

• Irreversible reaction

• Catalysed by pyruvate dehydrogenase

What is the structure of pyruvate dehydrogenase?

• Giant complex with a molecular mass of 4-10 million Da

• Made of 60 polypeptide chains

• Can be seen with electron microscopy

• Consists of 3 different enzymes

What are the three component enzymes of pyruvate dehydrogenase and what is their functions?

• E1 (24 chains)

• Catalyses the oxidative decarboxylation of pyruvate

• E2 (24 chains)

• Catalyses the transfer of the acetyl group to CoA

• E3 (16 chains)

• Catalyses the regeneration of the oxidised form of lipoamide

What is FAD and how is it helpful in metabolism?

• Redox-active coenzyme

• Major electron carrier in the oxidation of fuel molecules

• FAD + 2H+ + 2e- —> FADH2

• Forward reaction is reduction

• Backward reaction is oxidation

• Each FADH2 molecule is a carrier of 2 electrons

What happens during the first reaction of the citric acid cycle?

• Acetyl group is carried to the citric acid cycle by CoA

• Acetyl CoA (2C) combines with oxaloacetate (4C) to form citrate (6C)

• Catalysed by citrate synthase

• Uses H2O and releases the CoA to be reused

What happens during the second reaction of the citric acid cycle?

• Isomerisation of citrate (6C) to isocitrate (6C)

• Catalysed by aconitase in 2 steps

• Dehydration of citrate to cis-aconitate (intermediate)

• Hydration of cis-aconitate to isocitrate

What happens during reaction 3 of the citric acid cycle?

• Oxidation and decarboxylation of isocitrate (6C) to a-ketoglutarate (5C)

• Catalysed by isocitrate dehydrogenase in a two-step process

• Oxidation of isocitrate to oxalosuccinate (intermediate)

• Decarboxylation of oxalosuccinate to a-ketoglutarate

• 2 electrons and a proton are used to reduce NAD+ to NADH

What happens during the fourth reaction in the citric acid cycle?

• Oxidation and decarboxylation of a-ketoglutarate (5C) to succinyl CoA (4C)

• Catalysed by a-ketoglutarate dehydrogenase complex

• Two electrons and a proton are used to reduce NAD+ to NADH

• CoA is required in the reaction - similar to pyruvate dehydrogenase

What happens during the fifth reaction in the citric acid cycle?

• Cleavage of succinyl CoA (4C) into succinate (4C)

• High energy thioester bond is cleaved by succinyl CoA synthetase

• Release of CoA is coupled to phosphorylation of GDP/ADP to GTP/ATP - only ATP produced in the cycle

What happens during the sixth reaction in the citric acid cycle?

• Oxidation of succinate (4C) to fumarate (4C)

• Catalysed by succinate dehydrogenase

• Two electrons and two protons are transferred to FAD forming FADH2

What happens during the seventh reaction in the citric acid cycle?

• Hydrolysis/hydration of fumarate (4C) to malate (4C)

• Catalysed by fumarase

• Reaction requires the addition of H2O

What happens during the eighth reaction of the citric acid cycle?

• Oxidation of malate (4C) to oxaloacetate (4C)

• Catalysed by malate dehydrogenase

• Third and final production of NADH

What are the reagents put into one turn of the citric acid cycle?

• Acetyl CoA

• 3NAD+

• FAD

• GDP/ADP

• Pi

• 2H2O

What are the products formed from one turn of the citric acid cycle?

• 2CO2

• 3NADH

• FADH2

• GTP/ATP

• 2H+

• CoA

How many times does the cycle occur for a molecule of glucose?

• One molecule of glucose forms 2 acetyl CoA

• Cycle turns twice

• Forms 2ATP, 6NADH, 2FADH2 per molecule of glucose

What are metabolically important products made by the citric acid cycle?

• Convergent start point for carbohydrate, amino acid and fatty acid metabolism

• Citrate starts formation of fatty acids and sterols

• a-ketoglutarate starts formation of building blocks of amino acids and nucleotides

• Succinyl CoA starts formation of building blocks of porphyrins (eg haem)

What are the key enzymes of the citric acid cycle and how are they inhibited/activated?

• Pyruvate dehydrogenase - irreversible reaction

• Inhibited by ATP, Acetyl CoA and NADH

• Isocitrate dehydrogenase

• Inhibited by ATP and NADH

• Stimulated by ADP

• a-ketoglutarate dehydrogenase - rate limiting step

• Inhibited by ATP, succinyl CoA and NADH