Krebs cycle Energy - 215

Petr Tůma

Eva Samcová

Overview of Citric Acid Cycle

Key Concepts • The citric acid cycle (Krebs cycle) is a multistep catalytic process that converts acetyl groups derived from carbohydrates, fatty acids, and amino acids to CO2, and produces NADH, FADH2, and GTP.

Overview: Oxidative Fuel Metabolism

Reactions of Citric Acid Cycle

Tricarboxylic acid cycle (TCA)= Krebs

cycle – the hub of the metabolic system

• It accounts for the major portion of carbohydrate, fatty acid, amino acid oxidation and generates numerous biosynthetic precursors

• TCA is therefore amphibolic, that is, it operates both catabolically (biosynthesis of ATP) and anabolically (biosynthesis of amino acids, heme, glucose, lipids)

Starting compounds

• Acetyl-coenzyme A (acetyl-CoA)

• Oxaloacetate

• Cataplerotic reactions which utilize and therefore drain TCA

• Anaplerotic reactions replenish citric acid cycle intermediates

General features of CAC

• Oxidizes the acetyl-CoA not only from pyruvate

• The net reaction :3NAD+ + FAD + HS-CoA + GDP + Pi

→ 3NADH + H+ + FADH2 + GTP + CoA + 2CO2

● All enzymes of CAC are in mitochondrion, so all

substrates including NAD+ and GDP must be generated in

the mitochondrion or be transported to mitochondrion from

cytosol. Similarly all the products of CAC must be

consumed in mitochondrion or transferred to cytosol.

• The net effect of each round of CAC is oxidation of 1

acetyl group to 2CO2

Synthesis of Acetyl-Coenzyme A

Key Concepts • Pyruvate dehydrogenase is a multienzyme complex that catalyzes a five-part reaction in which pyruvate releases CO2 and the remaining acetyl group becomes linked to coenzyme A. • The reaction sequence requires the cofactors TPP, lipoamide, coenzyme A, FAD, and NAD+.

Cofactors of PDH Complex

Enzymes of the Citric Acid Cycle

Key Concepts • The eight enzymes of the citric acid cycle catalyze condensation, isomerization, oxidation–reduction, phosphorylation, and hydration reactions. • Two reactions produce CO2, one reaction produces GTP, and four reactions generate the reduced coenzymes NADH or FADH2.

Reaction 2: Aconitase

Reaction 3: Isocitrate Dehydrogenase

Releases 1st CO2

Reaction 4: α-Ketoglutarate

Dehydrogenase Resembles PDH

Complex—2nd NADH

Reaction 6: Succinate Dehydrogenase

Produces FADH2

Malonate Competitively Inhibits

Succinate Dehydrogenase

Reaction 7: Fumarase

Reaction 8: Malate Dehydrogenase

Produces 3rd NADH

Products of Citric Acid Cycle

Energy review

Reaction Cofactor ATP

isocitrate → 2-oxoglutarate NADH + H+ 3 (2,5)

2-oxoglutarate → succinyl-

CoA

NADH + H+ 3 (2,5)

succinyl-CoA → succinate - 1

succinate → fumarate FADH2 2 (1,5)

malate → oxaloacetate NADH + H+ 3 (2,5)

12 (10)

acetyl-CoA + 3NAD+ + Q + GDP + P + H2O →

→ 2CO2 + 3NADH+H+ + FADH2 + GTP +CoA

Regulation of Citric Acid Cycle

Regulation of TCA cycle activation – low rate of ATP/ADP and NADH/NAD+

inhibition – high rate of ATP/ADP and NADH/NAD+

The flux controlling enzymes

– Citrate synthase

– Isocitrate dehydrogenase

– 2-ketoglutarate dehydrogenase

• specific inhibitors

– fluoroacetate – aconitase

– Arsenic compounds inactivates lipoamide enzymes

(pyruvate dehydrogenase and 2-ketoglutarate

dehydrogenase

– malonate – succinate dehydrogenase

3 Enzymes Function Far From

Equilibrium

Amphibolic Functions of CAC

Metabolic Sources of Acetyl-CoA

• Acetyl-CoA (high energy compound)

the common product of carbohydrate,

fatty acids, and amino acids breakdown

• Pyruvate Dehydrogenase reaction

– pyruvate + NAD+ + CoA → acetyl-CoA + NADH/H+

+ CO2

– Irreversible reaction

– CoA, thiamine diphosphate, lipoamide, FAD, NAD+

• β-oxidation of fatty acids

• Amino acids (ketogenic AA, glucogenic AA)

Anabolic

reactions • citrate → fatty acids,

steroids

• 2-oxoglutarate → AAs of glutamate family

• succinyl-CoA → heme

• malate, oxaloacetate → glucose, AAs of pyruvate family

Anaplerotic Reactions

Replenish CAC Intermediates

Anaplerotic reactions replenishment of intermediates of KC

• pyruvate + CO2 + ATP → OAA + ADP – Pyruvate carboxylase

• Amino acids – Asp, Asn – OAA

– Glu, Gln, His, Pro, Arg – 2-oxoglutarate

– Ile, Val, Met, Trp – succinyl-CoA

– Ala, Ser, Thr, Cys, Gly - pyruvate

• Degradation of fatty acids with uneven (odd) number of carbon atoms:

Propionyl-CoA → succinyl-CoA

• Acetyl-CoA has no anaplerotic effect

The Mitochondrion

• The site of eukaryotic oxidative

metabolism with enzymes that mediate

this process

• Is bounded by a smooth outer membrane

and contains invaginated inner membrane

• The inner mitochondrial membrane is

impermeable to most hydrophilic

compounds therefore contain specific

transport systems (proteins 80%)

Mitochondrion - metabolic functions

Mitochondrion - structure

Transport of substances through

inner mitochondrial membrane

Shuttles Transfer of NADH + H+ from

cytoplasm into mitochondrion

1. The malate-aspartate shuttle

heart, liver, kidney

2. The glycerolphosphate shutles

– Glycerol-3-phosphate /Dihydroxyacetone phosphate

– brain, skeletal muscles

The malate-aspartate shuttle

The glycerolphosphate shuttle