Cell Bio: Chapter 12- Krebs Cycle and ETC

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Three parts of the mitochondria

Inner Membrane (phospholipid bilayer): has pyruvate transporters

Intermembrane Space: Collects H+

Outer Membrane (phospholipid bilayer): fairly permeable to most molecules

Pyruvate Dehydrogenase/Decarboxylase

• Converts pyruvate into an acetyl group thru oxidative decarboxylation

  • Pyruvate is oxidized (electrons are given to NAD+ —> NADH)

  • CO2 is released

• Acetyl group (2 carbon molecule) is linked to Coenzyme A (temporary carrier of acetyl group)

Parts of Coenzyme A

• Nucleotide (adenine, ribose, phosphate group)

• Connected to pantothenic acid (B vitamin) which contains a sulfhydryl group

  • Vitamins: cofactor for enzymes aka conenzymes (ex. NAD)

• Sulfhydryl group directly interacts with and binds to acetyl group made from pyruvate via PDH

Krebs Cycle/Citric Acid Cycle

Happens twice in the matrix of the mitochondria

1. Citrate Synthase adds acetyl-CoA to oxaloacetate to form citric acid (citrate which is in its anion form)

2. Aconitase converts citrate into isocitrate (isomerization reaction)

3. Isocitrate dehydrogenase oxidize isocitrate and remove (2) CO2 to form alpha ketoglutarate

   1. Electrons are given to NAD+ to form NADH (2)

4. Alpha ketoglutarate dehydrogenase oxidizes and decarboxyliases alpha ketoglutarate to form a succinyl group which linked to CoA to form succinyl-CoA

5. Succinyl-CoA synthetase takes phosphate off succinyl-CoA and gives it to GDP to form GTP which forms succinate

   1. GTP (energetically equivalent to ATP) is formed via substrate level phosphorylation

   2. Bond between succinyl group and CoA is high energy so it can allow for substrate level phosphorylation

6. Succinate dehydrogenase oxidizes succinate to form fumarate

   1. Uses FAD as temporary electron carrier that can reduced into FADH2

      1. Electrons come from a C-H bond

7. Fumarate Hydratase converts fumarate into malate

8. Malate dehydrogenase oxidizes malate to form oxaloacetate

   1. Uses NAD as temporary electron carrier because electrons come from C=O

Flavin Adenine Dinucleotide (FAD)

• Contains adenine, ribose, pyrophosphate bridge, and riboflavin (nitrogenous base and B vitamin)

• Riboflavin like nicotinamide can easily pick up electrons and become reduced

Redox Potential

Oxidation of different bonds yields electrons with different levels of energy

FAD and NAD have different redox potentials

Why is FAD used as an electron carrier for succinate instead of NAD?

• Electrons can only be transferred to a molecule with an equal or lower redox potential

• NAD+ has a higher redox potential than the electrons that come from succinate oxidation so it CAN’T take those electrons

• FAD has a lower redox potential than the electrons that come from succinate oxidation so it CAN take those electrons

How are the dehydrogenases regulated?

High conc. of NADH then activity of these enzymes slows down

Most important regulatory step in Krebs Cycle

Technically not in the Krebs Cycle but before it

Pyruvate Dehydrogenase regulates the Krebs and is most active in its dephosphorylated form

• Negatively regulated by acetyl-CoA, NADH, ATP (when these are high in concentration, lower PDH activity

• Positively regulated by CoA, NAD+, AMP

Enzyme 2 (E2) and Enzyme 1 (E1) in Regulating Krebs Cycle

E2 = PDH Kinase: phosphorylates PDH

• High ATP/ADP ratio —> activity of E2 goes up because PDH can slow down

E1 = PDH phosphatase: dephosphorylates PDH

• Low ATP/ADP ratio —> activity of E1 goes up because PDH can speed up

Overall Equation and Where did all the energy go?

Glucose + 10 NAD+ + 2FAD + 4ADP + 4Pi —> 6CO2 + 10 NADH + 10 H+ + 2 FADH2 + 4 ATP

All the energy is stored in the temporary electron carriers (NAD+ & FAD)

Next Steps after Krebs Cycle (Aerobic Metabolism)

• NADH and FADH2 deliver electrons to ETC in inner membrane

• Electrons are given to oxygen (final electron acceptor) to make H2O

• ETC uses some of the energy to create a H+ gradient that is used to make ATP

ETC

• Consists of four different complexes that contain multiple electron carriers which include:

  • Flavoproteins

  • Cytochromes

  • Coenzyme Q

  • Iron-sulfur proteins

Flavoproteins

• Derived from Flavin

• Ex: FAD, Flavin mononucleotide (FMN)

• Located in the first complex and accepts electrons from NADH

Cytochromes

• Located in complex 3 & 4

• Temporary electron carrier using heme groups (contain iron in the center)

Conenzyme Q

• Temporary electron carrier that delivers electron from complex 1 to complex 3 and electrons from complex 2 to complex 3

• Soluble in membrane

Iron Sulfur Proteins

• Temporary electron carriers present in complexes 1, 2, and 3

  • Complex 1: accepts electrons from FMN

  • Complex 2: accepts electrons from FADH2

  • Complex 3: accepts electrons from Co-QH2 (reduced form of Co-Q)

Oxidation of NADH

ΔG°’ = -52.5 kcal/mol

Redox Potential of ETC components

• As you go down electron gradient, the lower the redox potential (electrons can only be accepted by a carrier that has a lower redox potential)

  • Some energy is lost to heat

  • Most energy is used to transport H+

How does NADH transferring its electrons power the proton gradient?

• Electrons from NADH are transported from complex 1 to 2 and 3 and that generates energy

• That energy pumps H+ from the matrix into the intermembrane space creating a gradient

Oxidation of FADH2

ΔG°’ = -45.9 kcal/mol

How does FADH2 transferring its electrons power the proton gradient?

• Electrons from FADH2 are dropped off at Complex 2

• Those electrons travel through complex 3 and 4 via Co-Q and cytochromes

• This powers an H+ gradient

Final Step of Cell Respiration

• H+ ions go through ATP synthase in the inner membrane

  • Cells use oxygen to help synthesize ATP in a process called chemiosmosis