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Lecture 10: Cellular Respiration — Glycolysis and the Citric Acid Cycle

General Biology I — Molecular & Cellular

Learning Objectives

By the end of this lecture, students will be able to:

1. Write the overall equation for aerobic cellular respiration
2. Describe the stages of cellular respiration and where each occurs in the cell
3. Trace the steps of glycolysis and identify key intermediates, enzymes, and energy yields
4. Explain the role of pyruvate oxidation in linking glycolysis to the citric acid cycle
5. Describe the steps of the citric acid cycle and its energy outputs
6. Explain what happens during fermentation when oxygen is absent

Lecture Content

I. Overview of Cellular Respiration

Cellular respiration is the metabolic process by which cells extract energy from organic molecules and convert it into ATP, the universal energy currency. The overall equation--C6H12O6 + 6O2 -> 6CO2 + 6H2O + energy (ATP + heat)--summarizes what is in reality an elaborate sequence of enzyme-catalyzed steps, with a Delta G of -686 kcal/mol. This energy is not released in a single explosive burst; instead, it is harvested in controlled increments through a series of redox reactions. In each such reaction, electrons are transferred from one molecule to another: oxidation is the loss of electrons (often accompanied by loss of hydrogen atoms), and reduction is the gain of electrons. The mnemonic OIL RIG--Oxidation Is Loss, Reduction Is Gain--helps keep these straight. In cellular respiration, glucose is progressively oxidized while oxygen serves as the final electron acceptor and is reduced to water.

The electrons liberated from glucose do not travel directly to oxygen. Instead, they are captured by electron carriers--primarily NAD+, which is reduced to NADH, and FAD, which is reduced to FADH2. These carriers then deliver their high-energy electrons to the electron transport chain, where the energy is used to produce the bulk of the cell's ATP.

Aerobic respiration proceeds through four stages: glycolysis (in the cytoplasm), pyruvate oxidation (in the mitochondrial matrix), the citric acid cycle (also called the Krebs cycle or TCA cycle, in the mitochondrial matrix), and oxidative phosphorylation (at the inner mitochondrial membrane).

II. Glycolysis

Glycolysis--literally "glucose splitting"--takes place in the cytoplasm and does not require oxygen, making it the universal starting point for both aerobic and anaerobic energy extraction. Through ten enzymatic steps, one molecule of glucose (6 carbons) is converted into two molecules of pyruvate (3 carbons each). The pathway is divided into two phases.

A. Energy Investment Phase (Steps 1-5)

During the first five steps, the cell spends two molecules of ATP to phosphorylate glucose and rearrange it into a form that can be cleaved. In step 1, hexokinase transfers a phosphate from ATP to glucose, producing glucose-6-phosphate. This is an irreversible, committed step--once phosphorylated, glucose is trapped inside the cell. In step 3, phosphofructokinase-1 (PFK-1) catalyzes the most critical regulatory reaction in glycolysis, phosphorylating fructose-6-phosphate to fructose-1,6-bisphosphate. PFK-1 is the rate-limiting enzyme and the primary point of metabolic control: it is allosterically activated by AMP, ADP, and fructose-2,6-bisphosphate (signals of low energy status) and inhibited by ATP and citrate (signals of energy abundance). In step 4, aldolase cleaves the six-carbon fructose-1,6-bisphosphate into two three-carbon molecules: glyceraldehyde-3-phosphate (G3P) and dihydroxyacetone phosphate (DHAP). DHAP is promptly isomerized to G3P in step 5, so two molecules of G3P continue through the payoff phase.

B. Energy Payoff Phase (Steps 6-10)

The payoff phase occurs twice per glucose molecule, once for each G3P. In step 6, G3P is oxidized and NAD+ is reduced to NADH, while an inorganic phosphate is incorporated. Step 7 generates ATP through substrate-level phosphorylation, a direct transfer of a phosphate group from the substrate to ADP. Step 10, catalyzed by pyruvate kinase, produces another ATP by substrate-level phosphorylation and yields pyruvate. Pyruvate kinase is itself regulated: it is activated by fructose-1,6-bisphosphate (a feed-forward signal) and inhibited by ATP and alanine.

C. Net Yield of Glycolysis (per glucose)

The net products are 2 ATP (four produced minus two consumed), 2 NADH, and 2 pyruvate. Though modest compared to the total ATP yield of aerobic respiration, glycolysis is vitally important: it is the only source of ATP for cells that lack mitochondria (such as mature red blood cells) and provides emergency energy when oxygen supply is limited.

<image>A flowchart of glycolysis divided into two colored sections. Top section (red, "Energy Investment Phase"): Glucose is phosphorylated by hexokinase (ATP consumed), converted through intermediates to fructose-1,6-bisphosphate by PFK-1 (ATP consumed), then cleaved by aldolase into two G3P molecules. Bottom section (green, "Energy Payoff Phase"): Each G3P is oxidized (NAD+ reduced to NADH), phosphorylated, and converted to pyruvate through substrate-level phosphorylation (2 ATP produced per G3P). Net yield box shows: 2 ATP, 2 NADH, 2 pyruvate. Key regulatory enzymes (hexokinase, PFK-1, pyruvate kinase) are highlighted with their activators and inhibitors listed.</image>

III. Pyruvate Oxidation (The Transition Reaction)

Before entering the citric acid cycle, pyruvate must be transported from the cytoplasm into the mitochondrial matrix via a specific transporter in the inner mitochondrial membrane. There, the pyruvate dehydrogenase complex--a massive multienzyme assembly requiring five coenzymes (NAD+, CoA, FAD, thiamine pyrophosphate, and lipoic acid)--catalyzes an irreversible oxidative decarboxylation. The three-carbon pyruvate loses one carbon as CO2, is oxidized (reducing NAD+ to NADH), and the remaining two-carbon acetyl group is attached to coenzyme A to form acetyl-CoA. Per glucose molecule, this transition produces 2 acetyl-CoA, 2 CO2, and 2 NADH. Pyruvate dehydrogenase is tightly regulated, inhibited by its products (acetyl-CoA, NADH, and ATP) and activated by its substrates (NAD+, CoA, ADP) and by calcium ions.

IV. The Citric Acid Cycle (Krebs Cycle / TCA Cycle)

The citric acid cycle is a cyclical pathway in the mitochondrial matrix that completes the oxidation of the acetyl group from acetyl-CoA. The two-carbon acetyl group combines with the four-carbon oxaloacetate to form the six-carbon citrate. Over eight enzymatic steps, citrate is progressively oxidized, releasing two molecules of CO2 and regenerating oxaloacetate to begin the cycle anew.

Each turn of the cycle produces 3 NADH, 1 FADH2, 1 GTP (equivalent to 1 ATP, generated by substrate-level phosphorylation at the succinyl-CoA synthetase step), and 2 CO2. Three enzymes are particularly important regulatory points: citrate synthase (the entry step, which is irreversible), isocitrate dehydrogenase (the rate-limiting step, activated by ADP and inhibited by ATP and NADH), and alpha-ketoglutarate dehydrogenase (structurally similar to the pyruvate dehydrogenase complex, inhibited by succinyl-CoA and NADH, and activated by calcium). Notably, succinate dehydrogenase (step 6) is the only enzyme of the cycle that is embedded in the inner mitochondrial membrane, where it also functions as Complex II of the electron transport chain.

Since each glucose molecule yields two acetyl-CoA molecules, the cycle turns twice per glucose, generating a total of 6 NADH, 2 FADH2, 2 GTP, and 4 CO2.

<image>A circular diagram of the citric acid cycle. Acetyl-CoA enters at the top, combining with oxaloacetate (4C) to form citrate (6C). The cycle proceeds clockwise through isocitrate, alpha-ketoglutarate (5C, first CO2 released), succinyl-CoA (4C, second CO2 released), succinate, fumarate, malate, and back to oxaloacetate. At each oxidation step, the electron carrier produced is shown (NADH or FADH2). GTP production at the succinyl-CoA to succinate step is indicated. Key regulatory enzymes are highlighted in boxes. The carbon count (6C, 5C, 4C) is noted at each intermediate.</image>

V. Total Energy Harvest from Glycolysis through TCA (per glucose)

At this point, only 4 ATP have been produced directly by substrate-level phosphorylation. The real payoff comes next: the 10 NADH and 2 FADH2 carry high-energy electrons to the electron transport chain, where the vast majority of ATP will be synthesized. Each NADH yields approximately 2.5 ATP and each FADH2 yields approximately 1.5 ATP, as will be detailed in Lecture 11.

VI. Fermentation (Anaerobic Pathways)

When oxygen is unavailable, the electron transport chain cannot operate and NAD+ cannot be regenerated by oxidative phosphorylation. Without NAD+, glycolysis grinds to a halt. Fermentation solves this problem by providing an alternative route to regenerate NAD+ from NADH, allowing glycolysis to continue producing ATP even in the absence of oxygen.

Lactic acid fermentation converts pyruvate directly to lactate, oxidizing NADH back to NAD+ in the process. This pathway operates in muscle cells during intense exercise (when oxygen delivery cannot keep pace with demand), in red blood cells (which lack mitochondria entirely), and in certain bacteria responsible for producing yogurt and sauerkraut. The lactate generated in muscles can be transported to the liver and converted back to glucose via the Cori cycle.

Alcohol (ethanol) fermentation proceeds in two steps: pyruvate is first decarboxylated to acetaldehyde (releasing CO2), and acetaldehyde is then reduced to ethanol while NADH is oxidized to NAD+. This pathway operates in yeast and some bacteria and is the biochemical basis of brewing, winemaking, and bread-making--the CO2 released during fermentation is what causes bread dough to rise.

Both forms of fermentation yield only 2 ATP per glucose (from glycolysis alone), making them far less efficient than aerobic respiration, which produces approximately 30-32 ATP. Obligate anaerobes rely entirely on fermentation and are actually killed by oxygen. Facultative anaerobes, such as yeast and E. coli, can switch between aerobic respiration and fermentation depending on oxygen availability, choosing the more efficient aerobic pathway when possible.

<image>A branching pathway diagram showing the fate of pyruvate. From glycolysis, pyruvate sits at a decision point. Left branch (aerobic, O2 present): pyruvate enters the mitochondrion for pyruvate oxidation and the citric acid cycle, leading to the ETC and ~30-32 ATP total. Right branch (anaerobic, no O2): splits into two sub-branches — lactic acid fermentation (pyruvate reduced to lactate, NAD+ regenerated; occurs in muscle cells) and alcohol fermentation (pyruvate decarboxylated to acetaldehyde then reduced to ethanol + CO2, NAD+ regenerated; occurs in yeast). Both fermentation pathways yield only 2 ATP net.</image>

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