BIO 121 Chapter 9

Metabolic pathways

Metabolic pathways

• harvest energy from high-energy molecules such as glucose

• cellular respiration is critical and often interacts with other pathways

• comprises thousands of different chemical reactions that may be organized and regulated

Cellular respiration

• occurs through a long series of carefully controlled redox reactions (conserves energy/prevents mini explosions) that use the electrons of high-energy molecules to make ATP

• oxygen atoms are reduced to form water

• glucose + 6 oxygen gas + ADP + inorganic Phosphate —> 6 carbon dioxide + 6 water + ATP

• consists of 4 processes: glycolysis, pyruvate processing, Krebs cycle, and electron transport and oxidative phosphorylation

Glycolysis overview

• a series of 10 reactions that occurs in the cytosol of eukaryotes and prokaryotes

• net yield of 2 NADH, 2 ATP, 2 H2O, and 2 pyruvate for every glucose

• glucose + 2 ATP —> 2 (NADH + H+) + 4 ATP + 2 Pyruvate + 2 H2O

Phosphofructokinase (PFK)

• uses ATP to phosphorylate the end of fructose-6-phosphate to form fructose-1,6-bisphosphate

• increases potential energy

Pyruvate processing

• occurs in the matrix of the mitochondria or the cytosol of prokaryotes

• for eukaryotes, pyruvate is transported from the cytosol to the mitochondrial matrix

• catalyzed by pyruvate dehydrogenase, an enormous enzyme complex which is regulated by a negative feedback loop involving ATP

• 2 pyruvate + 2 NAD+ + 2 Coenzyme A —> 2 acetyl CoA + 2 CO2 + 2 NADH

• Decarboxylation: the carboxyl group on pyruvate (the 3rd carbon) is released as CO2

Kinase

• enzyme that catalyzes the transfer of a phosphate group from ATP to another molecule

Phosphotase

• enzyme that removes phosphate group

Krebs cycle (citric acid cycle)

• also occurs in the matrix of the mitochondria or the cytosol of prokaryotes

• 2 turns of the citric acid cycle for each glucose molecule

• Potential energy is released to reduce coenzymes

• The acetyl group (2C) from acetyl CoA is transferred to oxaloacetate (4C) to form citrate (6C); oxaloacetate is regenerated at the end (cycle)

• 8 reactions

• 2 acetyl coA —> 6 NADH + 6H+ + 2 FADH2 + 2 ATP + 4CO2

Electron transport and oxidative phosphorylation

• electron transport chain consisting of 4 main protein complexes establishes a proton gradient that is used to produce ATP

• uses NADH and FADH2 produced in previous steps to generate the protein gradient, which contributes to the phosphorylation of ADP

• uses O2 (oxygen gas) and produces ATP and water

• occurs across the inner membrane of the mitochondria or the plasma membrane + the periplasm of prokaryotes

• a small amount of energy is released in each reaction; each successive bond/molecule in the ETC holds less potential energy ; after the ETC, most of the chemical energy from glucose is accounted for by a proton electrochemical gradient

• primary goal: make ATP

• secondary goal: regenerate NAD+

Periplasm

• space between the cell wall and the plasma membrane

2 fundamental requirements of cellular life

• energy to generate ATP

• a source of carbon to use as raw materials for synthesizing macromolecules

Catabolic pathways

• involve the breakdown of molecules

• often harvest stored chemical energy to produce ATP

Anabolic pathways

• result in the synthesis of larger molecules from smaller components

• often use energy in the form of ATP

Homeostasis

• maintenance of a stable internal environment under different environmental conditions

Energy investment phase (glycolysis)

• reactions 1 through 5

• uses 2 ATP molecules

• regulation of the metabolic pathway occurs during this phase (reaction 3, regulation of phosphofructokinase)

Energy payoff phase (glycolysis)

• reactions 6 through 10

• NADH is made and ATP is produced by substrate-level phosphorylation

Substrate-level phosphorylation

• 1 way to make ATP

• the ONLY way to produce ATP through glycolysis

• enzyme facilitates the transfer of a phosphate group from a substrate to ADP

Glycolysis regulation

• regulated by feedback inhibition

• high levels of ATP inhibit the third enzyme/step of glycolysis (phosphofructokinase), which have two binding sites for ATP

• when ATP binds to the regulatory site of phosphofructokinase, the reaction rate slows dramatically

Regulation of pyruvate processing

• when products of glycolysis and pyruvate processing are abundant, pyruvate dehydrogenase is phosphorylated, inducing a conformational change in the enzyme and inhibiting its activity

Citric acid cycle regulation

• can be turned off at multiple points via several different mechanisms of feedback inhibitions

• regulated at steps 1, 3, and 4 by ATP and NADH

• reaction rates are high when ATP and NADH are scarce; rates are low when ATP or NAHD are abundant

Oxidation of NADH and FADH2

• oxidized by membrane complexes

• NADH is oxidized when combined with the inner membrane of the mitochondria; in prokaryotes, it is oxidized by the plasma membrane

• molecules in the inner mitochondrial membrane can cycle between oxidized and reduced states

ETC Protein complexes

• most are composed of easily-oxidized proteins

• some accept only electrons, while others accept electrons plus protons; each complex has differing redox potentials

Ubiquinone (coenzyme Q, or simply Q)

• lipid-soluble, non-protein

• critical component of the ETC

• reduced by complexes I and II; moves throughout the hydrophobic interior of the electron transport chain membrane, where it is oxidized by complex III

Redox potential

• ability to accept electrons

• High positive value = more potential to GAIN electrons

• strong negative value = more potential to LOSE electrons

Complex I (ETC)

• NADH dehydrogenase oxidizes NADH

• transfers 2 electrons through proteins containing FMN prosthetic groups and Fe-S cofactors to reduce an oxidized form of Q

• 4 protons pumped out of the matrix to the intermembrane space per pair of electrons

Complex II (ETC)

• Succinate dehydrogenase oxidizes FADH2

• transfers the two electrons through proteins containing Fe-S cofactors to reduce an oxidized form of Q

• this complex is also used in step 6 of the Krebs cycle

• does not produce sufficient energy to pump protons

Complex III

• cytochrome c reductase oxidizes Q

• transfers 1 electron at a time through proteins containing heme prosthetic groups and Fe-S cofactors to reduce an oxidized form of cytochrome c

• 4 protons for each pair of electrons is transported from the matrix to the intermembrane space

Cyt c (cyctochrome c)

• reduced by accepting a single electron from complex III

• moves along the surface of the ETC membrane, where it is oxidized by complex IV

Complex IV

• cytochrome c oxidase oxidizes cyt c

• transfers each electron through proteins containing heme prosthetic groups to reduce oxygen gas, which picks up two protons from the matrix to produce water

• 2 additional protons are pumped out of the matrix of the intermembrane space

ATP Synthesis (ETC)

• fueled by chemiosmosis; uses the established proton gradient to create ATP using ATP synthase

ATP synthase

• located in the inner mitochondrial membrane in eukaryotes, or the plasma membrane in prokaryotes

• creates energy from the proton motive force of the proton gradient to chemical bond energy in ATP

• is a rotary machine that makes ATP as it spins

• consists of 2 components—an ATPase "knob"/F1 unit, and a membrane-bound, proton-transporting base/F0 unit, which is a rotor that turns as protons flow through it—that are connected by a shaft and held in place by a stator

• the spinning F0 unit changes the conformation of the F1 unit so that it phosphorylates ADP to form ATP

Oxidative phosphorylation

• oxidative = FADH2 and NADH are being oxidized

• phosphorylation = ADP —> ATP

• different from substrate-level phosphorylation because instead of potential energy activating the enzyme, kinetic energy activates the enzyme (movement of protons down their gradient)

• yields ~24-28 ATP per glucose

Chemiosmotic hypothesis

• the linkage between electron transport and ATP production by ATP synthase is indirect

• the synthesis of ATP only requires a proton gradient

Aerobic respiration

• O2, which has a very high redox potential, is the final electron acceptor

• most efficient—CO2 (single-carbon compound) is the byproduct

Anaerobic respiration

• some other compound is the final electron acceptor

• has a lower energy yield compared to aerobic respiration because oxygen is super electronegative and has a high redox potential

• less efficient—some other carbon-containing (organic) molecule is the byproduct (ethanol, lactic acid, etc)

• seen in some prokaryotes

Fermentation

• a metabolic pathway that regenerates NAD+ from NADH

• the electron in NADH is transferred to pyruvate

• serves as an emergency backup for aerobic respiration when there is not enough oxygen

• incomplete oxidation of glucose; much less efficient than cellular respiration

• produces 2 ATP per glucose, compared with about 29 ATP per glucose in cellular respiration

Lactic acid fermentation

• fermentation in which the product is lactic acid

• occurs in humans in the absence of oxygen

• muscle cramps = the accumulation of lactic acid

• in humans, lactic acid fermentation results in the production of yogurt, cheese, etc

• produces only 2 ATP (by substrate-level phosphorylation)

Ethanol fermentation

• some yeast cells can perform alcohol fermentation

• pyruvate is converted to acetaldehyde and CO2

• acetaldehyde accepts electrons from NADH

• ethanol and NAD+ are produced

Faculative anaerobes

• organisms that can switch between fermentation and aerobic respiration

• only use fermentation if an electron acceptor is not available

• E.coli, yeast, etc

Glycolysis step 1

• hexokinase uses ATP to phosphorylate glucose, increasing its potential energy

• forms glucose-6-phosphate and ADP

Glycolysis Step 2

• phosphoglucose isomerase converts glucose-6-phosphate to fructose-6-phosphate (an isomer)

Glycolysis Step 3

• Phosphofructokinase uses ATP to phosphorylate the opposite end of fructose-6-phosphate, increasing its potential energy

• forms fructose-1,6-bisphosphate

Glycolysis Step 4

• fructose-bis-phosphate aldolase cleaves fructose-1,6-bisphosphate into 2 different 3-carbon sugars (DAP and G3P)

Glycolysis Step 5

• triose phosphate isomerase converts dihydroxyacetone phosphate (DAP) to glyceraldehyde-3-phosphate (G3P)

• reaction is fully reversible, but DAP-to-G3P reaction is favored because G3P can be immediately used as a substrate for step 6

Glycolysis Step 6

• glyceraldehyde-3-phosphate (G3P) dehydrogenase catalyzes a 2-step reaction

• first oxidizes G3P using the NAD+ coenzyme to produce NADH

• Energy from this reaction is used to attach an inorganic phosphate to the oxidized product to form 1,3-bisphosphoglycerate

Glycolysis Step 7

• phosphoglycerate kinase transfers a phosphate from 1,3-bisphosphoglycerate to ADP to make 3-phosphoglycerate and ATP (PRODUCES ATP—1 for each 3-carbon intermediate)

Glycolysis Step 8

• phosphoglycerate mutase rearranges the phosphate in 3-phosphoglycerate to form 2-phosphoglycerate

Glycolysis Step 9

• enolase removes a water molecule from 2-phosphoglycerate to form a C=C double bond and produce phosphoenolpyruvate

Glycolysis Step 10

• remaining phosphate groups are added to 2 ADP molecules to form 2 ATP and pyruvate

• pyruvate kinase transfers a phosphate fro phosphoenolpyruvate to ADP to make pyruvate and ATP

Electron Transport Chain Theoretical Yield

• 1 NADH = 3 ATP

• 1 FADH2 = 2 ATP (lower because complex II, where FADH2 is oxidized, has a lower redox potential than complex I, where NADH is oxidized)

ETC Actual Yield

• 1 NADH = ~2.25 ATP

• 1 FADH2 = ~1.25 ATP