Citric acid cycle and respiratory chain

Pavla Balínová

MitochondriaStructure of mitochondria:• Outer membrane• Inner membrane (folded)• Matrix space (mtDNA, ribosomes, enzymes of CAC, β-oxidation of FA, heme synthesis,…)Function of mitochondria:• production of acetyl-CoA from pyruvate (PDH reaction)• production of ATP (by oxidative phosphorylation)• degradation of FA by β-oxidation • urea synthesis• heme synthesis,….

Citric acid cycle (CAC) tricarboxylic acid cycle, Krebs cycle

• CAC is a set of reactions which form a metabolic pathway for aerobic oxidation of saccharides, lipids and proteins.

• Reduced equivalents (NADH, FADH2) are released by sequential decarboxylations and oxidations of citric acid. These reduced equivalents are used to respiratory chain and oxidative phosphorylation to produce ATP

• CAC plays a key role in futher metabolic reactions (i. e. gluconeogenesis, transamination, deamination

or lipogenesis)

Function of CAC• Oxidation of CH3-CO- to 2 CO2 → formation of

reduced coenzymes NADH + H+ and FADH2

• CAC is a central junction of an intermediary metabolism = amphibolic pathway

→ catabolic pathways generate intermediates into CAC

→ anabolic pathways withdraw some intermediates from CAC (oxaloacetate → gluconeogenesis, succinyl-CoA → synthesis of porphyrins etc.)

Coenzyme A (CoA)

Figure was assumed from http://www.lipidlibrary.co.uk/Lipids/coa/index.htm

-CO-CH3 acetyl

Figure was assumed from http://www.biocarta.com/pathfiles/KrebPathway.asp

Result of CAC:Production of 2 mol CO2, 3 mol NADH + H+, 1 mol FADH2, 1 mol GTP

Anaplerotic (support) reactions:• Pyr + CO2 + ATP → oxaloacetate + ADP + Pi (pyruvate carboxylase)

• degradation of most amino acids gives the following intermediates of CAC: oxaloacetate, α-ketoglutarate, fumarate

• Propionyl-CoA → succinyl-CoA

Regulation of CACRegulatory factors of CAC are:• NADH / NAD+ ratio• ATP / AMP ratio• availability of CAC substrates and energy situation within the cell

Regulatory enzymes of CAC:Citrate synthase is mainly regulated with availability of acetyl-CoA and oxaloacetate.Isocitrate dehydrogenase and α-ketoglutarate dehydrogenase are inhibited by ↑ NADH / NAD+. On the contrary, these enzymes are activated by AMP and NAD+.

The activity of CAC is closely linked to the availability of O2.

Transport of acetyl-CoA within the cell

• acetyl-CoA + oxaloacetate → citrate (citrate synthase in CAC)

• citrate is exported from mitochondria to cytoplasm in exchange for malate

• citrate is cleaved to acetyl-CoA and oxaloacetate (citrate lyase) in the cytoplasm

• reduction of oxaloacetate to malate (malate dehydrogenase = „malic enzyme“ – production of NADPH + H+)

• malate is returned by antiport into mitochondria or it is oxidative decarboxylated to pyruvate

Respiratory chain Reduced coenzymes NADH and FADH2 release H atoms (e- and H+) in electron transport system.

•location: inner mitochondrial membrane

•composition: enzyme complexes I – IV, 2 mobile carriers of electrons – coenzyme Q (ubiquinone) and cytochrome c

•function: transport of electrons in series of redox reactions and H+. Oxygen is a final acceptor of electrons.H+ are transmitted by complexes I, III and IV. Proton gradient is used to move ATP-synthase.

Figure was assumed from http://web.indstate.edu/thcme/mwking/oxidative-phosphorylation.html

Figure was assumed from http://www.biocarta.com/pathfiles/h_etcPathway.asp

Oxidative phosphorylation

H+ are ejected from mitochondrial matrix into intermembrane space by complexes I, III and IV.These protons create an electrochemical gradient across the inner mitochondrial membrane. Energy of this gradient is used for movement of ATP synthase. This enzyme allows protons to flow back down their concentration gradient across the membrane.

Figure was assumed from http://en.wikipedia.org/wiki/Electron_transport_chain

Respiratory chain + oxidative (aerobic) phosphorylation

Figure was assumed from http://www.rpi.edu/dept/bcbp/molbiochem/MBWeb/mb1/part2/oxphos.html

ATP-synthase

ATP-synthase consists of 2 subunits:F0 – in inner mitochondrial membrane (a proton channel)F1 – catalytic unit (matrix)Figure was assumed from http://en.wikipedia.org/wiki/ATP_synthase

Uncoupling proteinsUncoupling proteins (UCP) are compounds which allow protons to flow across the mitochondrial membrane with low production of ATP.Energy of proton gradient is released as heat.

UCP-1 (thermogenin) – brown adipose tissue (newborns)UCP-2 – mainly white adipose tissue UCP-3 – skeletal musclesUCP-4,5 – brainHibernating mammals

Uncoupling proteins (UCP)

UCP-1Thermogenin

Energy of proton gradient is released as heat.

Clinical correlation● Hypoxia= a lack of O2 in the inner mitochondrial membranecauses: lack of O2 in breathed air, myocardial infarction, anemia, atherosclerosisresult: failure of ATP formation

● Cyanide poisoningIon CN- binds to the complex IV (Fe3+ in heme of the cytochrome a) and blocks an electron transport to O2 → stopping of respiratory chain and synthesis of ATP → a rapid failure of cellular functions and death