Cellular Respiration

Cellular Respiration

Overview

• All living organisms must harvest energy from complex molecules to power cellular functions.
• Energy stored in covalent bonds is transferred to high-energy carriers.
• Cellular respiration is a highly controlled and regulated process.

Key Concepts

• Cellular respiration: Harvests chemical energy.
• Aerobic Respiration:
  • Cell Respiration
  • Glycolysis
  • Citric Acid Cycle
  • Oxidative Phosphorylation
• Anaerobic Respiration:
  • Fermentation
  • Glycolysis
  • Fermentation

Location and Products

• Glycolysis: Occurs in the cytosol, produces pyruvate and ATP.
• Citric Acid Cycle: Occurs in the mitochondria, produces ATP.
• Oxidative Phosphorylation: Occurs via electron transport and chemiosmosis.
• Fermentation

Energy Metabolism Control

• Oxidation in a series of steps allows the controlled release of energy, preventing it from being lost as heat.

Redox Reactions

• Redox Reactions: Transfer of electrons from one reactant to another.
• Oxidation: Loss of an electron (X is oxidized).
• Reduction: Gain of an electron (Y is reduced), reducing the positive charge.
• $Xe^- + Y \rightarrow X + Ye^-$
• X becomes oxidized (loses electron).
• Y becomes reduced (gains electron).
• Reducing Agent: Electron donor (X) gets oxidized.
• Oxidizing Agent: Electron acceptor (Y) gets reduced.
• Redox reactions are always coupled.

Relocation of Electrons

• Electrons lose potential energy when shifting from a less electronegative atom to a more electronegative one (e.g., H → O or C → O).
• This releases energy (exergonic, spontaneous).
• Losing potential energy is like water flowing downhill.
• Moving from high to low energy states (unstable to stable) releases energy.
• Hydrogen atoms are unstable and in a higher energy state, making it easy to pull electrons from them.
• Once bound to oxygen, electrons are hard to pull away due to oxygen's high electronegativity, resulting in a stable, lower energy state.

Organic Molecules as Fuels

• Organic molecules with abundant hydrogen atoms are excellent fuels.
• H-bonds are a source of hilltop electrons.
• Energy is released as electrons fall down the energy gradient when transferred to oxygen.
• Unstable → Stable
• High energy → Low energy
• Less electronegative → more electronegative
• Main energy foods (carbs & fats) are reservoirs of electrons associated with hydrogen.

Electron Transfer Mechanisms

• Reactions use H atoms to transfer electrons.
• Hydrogen atom: 1 proton + 1 electron.
• Proton: Hydrogen ion ($H^+$), stripped of its electron.
• Electrons are first transferred to NAD+ (coenzyme).
• NAD+: Nicotinamide adenine dinucleotide, a derivative of niacin (vitamin B).
• NAD+ is an oxidizing agent; it gets reduced (gains an electron).
• NAD+ is an electron shuttle, becoming NADH when it carries electrons.

NAD+/NADH

• NAD+ is reduced to NADH.
• Dehydrogenase: Enzyme that removes 2 hydrogen atoms from glucose (oxidizing glucose).
• Delivers one H atom and the electron from the 2nd H atom.
• $H-C-OH + NAD^+ \xrightarrow{\text{Dehydrogenase}} C=O + NADH + H^+$

ATP Production Mechanisms

1. Substrate-level phosphorylation: Direct transfer of energy to form a phosphodiester bond, forming ATP.
   • Occurs during glycolysis and the citric acid cycle.
   • Enzymes attach phosphate onto a metabolite.
2. Oxidative phosphorylation: Activated carriers transfer energy through indirect mechanisms to drive ATP production.
   • Responsible for the majority of ATP produced in aerobic respiration.
   • Involves the electron transport chain and chemiosmosis.

Complete Glucose Oxidation

• Many sources (protein, fats, carbs) feed into aerobic respiration through various pathways.
• Focus on glucose metabolism as it generates the most energy in many animal cells.
• Complete Oxidation of Glucose Occurs in 3 Stages:
  1. Glycolysis: Anaerobic breakdown of glucose into 2 pyruvate, generating net 2 ATP and 2 NADH.
  2. Citric Acid Cycle: Complete breakdown of acetyl-CoA, generating 2 GTP, 6 NADH, 2 FADH2.
  3. Oxidative Phosphorylation: Converts energy from electron carriers to ATP; majority of ATP synthesis.

Glycolysis

• $Glucose + 2 ATP \rightarrow 2 Pyruvate + 2 ATP + 2 NADH$
• 10-step process, each facilitated by a specific enzyme.
• Series of small rearrangements producing various sugar intermediates.
• Occurs in the cytoplasm.
• Anaerobic.

3 Phases of Glycolysis

1. Energy Investment:
   • 2 ATP are used in preparation for splitting into 2 molecules.
   • Step 1: Hexokinase
   • Step 3: Phosphofructokinase
2. Cleavage:
   • Splits 6-carbon glucose into (2) 3-carbon molecules.
3. Energy Generation:
   • Production of 4 ATP (steps 7 and 10) and 2 NADH (Step 6).

Regulatory Steps of Glycolysis

• Key regulatory enzymes include hexokinase, phosphoglucose isomerase, and phosphofructokinase.
• Phosphofructokinase is the most important regulatory step.
• ATP inhibits, while AMP activates.

Energy Investment Details

• In steps 1 and 3 of glycolysis, ATP is used to phosphorylate substrates.
• The addition of a phosphate group raises the product to a higher free energy level.
• This allows later endergonic steps to occur.
• Example free energy changes:
  • $\Delta G = -4 \text{ kcal/mol}$
  • $\Delta G = +0.4 \text{ kcal/mol}$
  • $\Delta G = -3.4 \text{ kcal/mol}$
  • $\Delta G = +5.7 \text{ kcal/mol}$
• Overall $\Delta G = -1.3 \text{ kcal/mol}$

NADH Production

• In step 6 of glycolysis, glyceraldehyde 3-phosphate dehydrogenase breaks a C-H bond on glyceraldehyde 3-phosphate.
• Transfers the H and another electron to NAD+ → NADH.
• NADH is carrying 2 electrons.
• Replaced by a phosphate group, creating a high-energy bond.
• Producing 1,3-biphosphoglycerate through substrate-level phosphorylation.

ATP Production Details

• Step 7: Phosphoglycerate kinase breaks the high-energy phosphate bond on 1,3-biphosphogyerate.
  • Transfers the phosphate group to ADP → ATP, resulting in 3-phosphoglycerate.
• Step 10: Pyruvate kinase transfers the remaining high-energy phosphate group from phosphoenolpyruvate to ADP → ATP.
  • High-energy bond formed during step 9.
  • Resulting in pyruvate, which will feed into fermentation OR aerobic respiration.

Fermentation

• Used by organisms that do NOT use oxygen to generate ATP (bacteria, yeast).
• They make all ATP through substrate-level phosphorylation in glycolysis (2 ATP total).
• Occurs in the cytosol.
• Glycolysis yields 2 ATP via substrate-level phosphorylation.
• Fermentation oxidizes NADH back into NAD+ to keep glycolysis going when there is no ETC to oxidize it back to NAD+.
• Primary purpose: Oxidation of NADH → NAD+.
• Replenishes cytosolic NAD+ to enable glycolysis to continue.
• Results in conversion of pyruvate to:
  • Lactate (muscle cells) OR
  • Ethanol + $CO_2$ (yeast).
• NO ENERGY PRODUCED in fermentation but REQUIRED for glycosylation to continue in the ABSENCE of oxygen!

Alcohol Fermentation

• Glucose undergoes glycolysis to produce 2 pyruvate, yielding 2 ATP and 2 NADH.
• 2 Pyruvate are converted to 2 Acetaldehyde, releasing $2CO_2$.
• 2 Acetaldehyde is reduced by 2 NADH to form 2 Ethanol, regenerating $2NAD^+$.

Lactic Acid Fermentation

• Glucose undergoes glycolysis to produce 2 pyruvate, yielding 2 ATP and 2 NADH.
• 2 Pyruvate is directly reduced by 2 NADH to form 2 Lactate, regenerating $2NAD^+$.

Aerobic Respiration

• Occurs in the mitochondria.

Mitochondrial Structure

• Outer membrane:
  • Porins – small molecule diffusion.
  • Translocase (TOM) – selective transport.
• Inner membrane (Cristae):
  • Translocase (TIM).
  • Oxidative Phosphorylation machinery
  • Electron Transport Chain.
  • ATP synthase.
• Intermembrane space:
  • High proton concentration (low pH).
• Matrix:
  • Many soluble enzymes.
  • Citric Acid Cycle.
  • High pH compared to intermembrane space.
  • Low proton [$H^+$].
• Cristae increase surface area.

Cellular Respiration Equation

• Complete oxidation of macromolecules through a series of chemical reactions takes place in the mitochondria.
• $C<em>6H</em>{12}O<em>6 + 6 O</em>2 \rightarrow 6 CO<em>2 + 6 H</em>2O + \text{Energy (ATP + heat)}$
• Simple, balanced equation – cellular process is much more complicated.

Link Reaction: Pyruvate to Acetyl-CoA

• Pyruvate is actively pumped into the mitochondrial matrix.
• Pyruvate dehydrogenase complex (complex of 3 enzymes):
  1. Decarboxylates pyruvate (removal of $CO_2$ = waste).
  2. Generates NADH (glucose is oxidized, $NAD^+$ is reduced).
  3. Acetyl group combines with Coenzyme A to produce Acetyl-CoA.
• Many other macromolecules are converted to acetyl-CoA in the mitochondrial matrix, including fatty acids and amino acids.

Citric Acid Cycle

• Complete oxidation of Acetyl-CoA occurs within the mitochondrial matrix.
• Indirect requirement of oxygen.
• Each Acetyl-CoA oxidized results in:
  • 1 GTP (equivalent to ATP).
  • 3 NADH.
  • 1 FADH2.
  • 2 $CO_2$
• Main contributing product = high energy electron carriers NADH and FADH2.
• Each glucose molecule yields 2 pyruvate → 2 Acetyl-CoA.
• 3 water molecules are required for each cycle.

Overview of Citric Acid Cycle

Steps:

• Acetyl CoA (2C) combines with oxaloacetate (4C) to form citrate (6C).
• Citrate is converted to isocitrate (6C).
• Release of $CO_2$ and production of NADH + $H^+$ converts isocitrate to \alpha-ketoglutarate (5C).
• Release of $CO_2$, production of NADH + $H^+$, and addition of HS-CoA converts \alpha-ketoglutarate to succinyl CoA (4C).
• Addition of GDP + Pi converts succinyl CoA to succinate (4C), producing GTP and HS-CoA.
• Production of FADH2 converts succinate to fumarate (4C).
• Addition of $H_2O$ converts fumarate to malate (4C).
• Production of NADH + $H^+$ converts malate to oxaloacetate (4C).

FADH2 Details

• Similar to NADH, binds 2 hydrogen atoms (= carrying 2 electrons).
• $FAD + 2H^+ + 2e^- \rightarrow FADH_2$

Oxidative Phosphorylation

• Electron Transport Chain and Chemiosmosis.

Two Processes

1. Electron transport chain (ETC): Sets up a $H^+$ gradient across the inner mitochondrial membrane.
   • Protein complexes pump $H^+$ into the intermembrane space.
   • High [$H^+$] in the intermembrane space.
   • Low [$H^+$] in the matrix.
2. Chemiosmosis: As $H^+$ diffuses from the intermembrane spaces, across the inner membrane into the matrix, they pass through ATP synthase, which catalyzes the formation of ATP.

Stages

1. Electron Transport Chain (ETC):
   • Conversion of energy from electron carriers to a proton electrochemical gradient across the inner mitochondrial membrane.
   • ETC embedded in the inner mitochondrial membrane.
2. Chemiosmosis:
   • Use of the proton gradient to power ATP synthase.
   • ATP synthase produces ATP.

• A proton is a hydrogen ion!

Electron Transport Chain

• Series of complexes embedded in the inner-membrane.
• As electrons are passed from one complex to the next, energy is released.
• Energy is used to pump protons across the inner-membrane, creating a high concentration of protons in the intermembrane space.
• No ATP is directly produced in the ETC but required to power ATP synthase.
• Main Objective: set up $H^+$ gradient!

ETC Complexes

• Arranged in Order of Increasing Redox Potential.
• Redox potential is a measure of affinity for electrons.
• Larger values indicate strong affinity – tendency to accept electrons.
• Smaller/negative values indicate weak affinity – tendency to donate electrons.
• Complexes are ordered in increasing affinity due to electron loss of energy as they move through the ETC.
• NADH dehydrogenase has a lower redox potential.
• Cytochrome c oxidase has a higher redox potential.
• $O_2$ is the most electronegative!

Action of ETC

• Actively pumping $H^+$ out of the matrix creates a high $H^+$ concentration in the intermembrane space.
• $H^+$ diffuses back into the matrix where there is a low $H^+$ concentration.

ETC Components

• NADH, FADH2, FMN, Fe•S, FAD, Q, Cyt b, Cyt c, Cyt c1, Cyt a, Cyt a3.
• Electrons are passed down the chain, each component becoming reduced when it accepts electrons and oxidized when it passes them on.
• $O_2$ is the final electron acceptor, combining with electrons and protons to form water.

Free Energy Change

• Each component of the chain becomes reduced when it accepts electrons from its uphill neighbor and becomes oxidized again as it passes the electron downhill.

Importance of Oxygen

• Without electronegative $O_2$ to pull electrons down the ETC, oxidative phosphorylation would stop.

ETC Functions

• Oxidizes NADH → $NAD^+$ and FADH2 → FAD.
• Generates a proton gradient.
• Forms water.
• $O<em>2$ is the final electron acceptor (4 electrons + 4 protons + $O</em>2$ → 2 $H_2O$).

ATP Synthase

• Multi-subunit complex with a rotor, stator, rod, and knob.
• $ADP + P_i \rightarrow ATP$

ATP Synthase Mechanism

• The rotor within the membrane spins as $H^+$ flows past it down the $H^+$ gradient.
• The stator, anchored in the membrane, holds the knob stationary.
• The rod, extending into the knob, also spins, activating catalytic sites in the knob.
• 3 catalytic sites in the stationary knob join inorganic phosphate to ADP to make ATP.

Efficiency

• About 50% of the energy in a glucose molecule is used to make ~30 ATP; the other energy is lost as heat.

Summary

• The whole point of cell respiration is to use glucose to make ATP, which drives all active cell processes.
• Electrons are removed from food to create a $H^+$ gradient, which makes ATP.
• Glycolysis: Function is to oxidize glucose (remove electrons and send them to ETC).
• $NAD^+$ is the electron shuttle, an oxidizing agent that oxidizes glucose and is reduced to NADH.
• Oxidation of Pyruvate: A 3-step process after glycolysis but before the citric acid cycle.
• Converts pyruvate into acetyl coenzyme A and creates $CO_2$.
• Citric Acid Cycle: Function is to complete the oxidation of pyruvate (actually acetyl coenzyme A).
• Removes electrons from acetyl coenzyme A and sends them to ETC.
• Uses $NAD^+$ AND FAD as e- shuttles.
• $NAD^+$ is reduced to NADH.
• FAD is reduced to FADH.
• Oxidative Phosphorylation: 2 processes
  • Electron Transport Chain: Function is to generate $H^+$ (proton) gradient across the inner mitochondrial membrane.
    • $H^+$ is high in the intermembrane space (acidic).
    • $H^+$ is low in the matrix (basic).
    • Oxygen is the terminal electron acceptor.
  • Chemiosmosis: Diffusion of $H^+$ from the intermembrane space into the matrix through ATP synthase, which generates a large amount of ATP.

Energy Balance

• Glycolysis: 2 ATP via substrate-level phosphorylation in the cytosol.
  • 2 pyruvates are the end product; most of the energy is still here.
• Oxidation of Pyruvate: 3-step process, $CO_2$ generated.
• Citric Acid Cycle: 2 ATP via substrate-level phosphorylation in the mitochondrial matrix.
  • Produces $CO_2$.
• Oxidative Phosphorylation: 28-30 ATP via chemiosmosis.
  • Mitochondrial inner membrane- ETC (consumes oxygen).
  • Chemiosmosis: Diffusion of $H^+$ from the intermembrane space into the matrix (crosses the inner mitochondrial membrane) through ATP synthase.

Redox Reactions

• Oxidized: Lose e- (reducing agent).
• Reduced: Gain e- (oxidizing agent).
• Example: Glucose reduces $NAD^+$. Glucose is the reducing agent and is oxidized. $NAD^+$ is reduced and is the oxidizing agent.
• Glucose is oxidized into pyruvate.
• $NAD^+$ is reduced into NADH.

Review Questions

• Explain the difference between substrate-level phosphorylation and oxidative phosphorylation.
• List the 3 major metabolic pathways that complete the oxidation of glucose.
• What are the 3 outputs of glycolysis? Where is most of the energy at the end of that pathway?
• Explain why 2 ATP are needed in glycolysis (what is the function of those phosphates?).
• Where does glycolysis occur? Is it aerobic or anaerobic?
• What is the function of NADH?
• What is the main function of glycolysis?
• Is fermentation aerobic or anaerobic? Where does it occur in the cell?
• What type of phosphorylation occurs in fermentation (substrate level or oxidative)?
• Explain why fermentation is necessary (what is the primary purpose?).
• What does the metabolic pathway of fermentation produce?
• Who is reduced and who is oxidized in alcohol fermentation?
• Who is reduced and who is oxidized in lactic acid fermentation?
• What is glycogen? How is it different from glucose? Where is it stored in the body?
• What is gluconeogenesis? When is this process utilized?
• What is meant by the term oxidation?
• Be familiar with the structure of the mitochondria as it is key to understanding the processes of oxidative phosphorylation.
• List the 3 steps of converting pyruvate to acetyl-CoA.
• What is produced in this process (list everything)?
• Where does it occur?
• Who is reduced? Who is oxidized?
• Where does the citric acid cycle occur? Be specific!
• What does 2 turns of the citric acid cycle produce?
• What is the function of FADH2?
• Is it higher or lower energy than NADH?
• Does the citric acid cycle use oxygen? Does it produce CO2?
• What is the main function of the citric acid cycle?
• Where does oxidative phosphorylation occur in the cell?
• Does it use oxygen? Does it produce CO2?
• What is the main function of the ETC?
• What is the main function of chemiosmosis?
• Explain what the proton motive force is in your own words.
• Explain how ATP synthase works.
• What molecule is the final electron acceptor of the ETC?
• What is the most important regulatory step in glycolysis?
• How is it regulated (what turns the enzyme on/off?)?