topic 10

A Western biochemistry club annual general meeting is scheduled for tonight at 2316. Information regarding midterm review sessions and writing locations will be posted soon. The midterm is on November 1st.

Topic 10: Metabolism

This topic covers mitochondrial structure, metabolic fates of pyruvate, the citric acid cycle, and its regulation relative to glycolysis and other inhibitors. It's important to understand how these pathways intersect.

Relationship to Previous Topics

Topic 9 covered glycogen synthesis and breakdown, glycolysis (glucose to pyruvate), fermentation (glucose to lactate), and gluconeogenesis (pyruvate back to glucose). The pentose phosphate pathway was also discussed. Topic 10 focuses on the fate of pyruvate after glycolysis.

Pyruvate's Role in Cellular Respiration

Pyruvate is converted to acetyl-CoA, which then enters the citric acid cycle. This process, also known as cellular or mitochondrial respiration, involves oxidizing carbon compounds (like pyruvate) to CO2CO_2 and using the process to generate ATP.

Mitochondrial Structure
Location of Processes

  • Glycolysis, Gluconeogenesis, Pentose Phosphate Pathway, Glycogen Storage: Occur in the cytosol.

  • Pyruvate Oxidation, Citric Acid Cycle: Occur in the mitochondrial matrix. Pyruvate must be transported into the mitochondria.

Key Features of Mitochondria

  • Membranes: Possess two distinct membranes: an outer membrane and an inner membrane.

  • Genome: Contains its own DNA, separate from nuclear DNA, encoding less than 40 genes. Mitochondria replicate by splitting, similar to bacteria.

  • Endosymbiotic Theory: This theory, championed by Lynn Margulis in the 1960s-70s, posits that mitochondria evolved from free-living bacteria that entered a symbiotic relationship with another cell. Genes were eventually transferred from mitochondria to the nucleus. Margulis's work highlights perseverance, as her ideas were initially rejected by many scientific journals.

Mitochondrial Compartments

  1. Outer Membrane:

    • Permeable to polar molecules up to approximately 5000 Daltons due to the presence of porins (beta-barrel structures).

    • Excludes larger molecules like proteins.

  2. Intermembrane Space:

    • The space between the outer and inner membranes.

    • Extends into the organelle due to folds in the inner membrane (cristae), which increase surface area and create microenvironments.

  3. Inner Membrane:

    • Not very permeable to small molecules; transport is tightly regulated and requires protein assistance for ions and small polar molecules.

    • Crucial for maintaining ion gradients (discussed in Topic 11).

  4. Matrix:

    • The space enclosed by the inner mitochondrial membrane.

    • Contains the enzymes for pyruvate oxidation and the citric acid cycle.

Coenzyme A (CoA)

  • Structure: Resembles an ADP molecule with an extra phosphate, but also includes a pantothenic acid residue (derived from Vitamin B5, an essential dietary vitamin).

  • Functional Group: The critical component is the thiol group (-SH).

    • This is the site of modification for Coenzyme A, often with fatty acid groups, forming fatty acyl groups.

    • Acetyl-CoA is a classic example, where a two-carbon acetate group (a short fatty acid) is joined to the thiol group via a thioester linkage.

  • Thioester Linkage: This is considered a "high-energy bond." It means that the group is activated, and reactions involving its cleavage will have a negative ΔG\Delta G (favorable process), releasing free energy.

  • Function: Coenzyme A acts as a shuttle, picking up and dropping off fatty acid groups in various metabolic pathways and activating them for subsequent reactions.

  • Discovery: Discovered by Fritz Lipmann, who received the Nobel Prize in 1953. Lipmann also suggested that ATP is a primary energy source for many reactions and coined the term "high-energy bond."

Pyruvate Oxidation

  • Location: Mitochondrial matrix.

  • Enzyme: Pyruvate dehydrogenase complex. This is a very large and complicated enzyme consisting of dozens of polypeptide chains and three distinct enzymes. The specific chemistry is not detailed but the overall reaction is important.

  • Reaction:

    • Pyruvate (3 carbons) undergoes oxidative decarboxylation.

    • One carbon is oxidized to CO2CO_2 (exhaled).

    • The remaining two carbons form an acetyl group, which is attached to Coenzyme A, forming acetyl-CoA.

    • Electrons removed during oxidation are transferred to NAD+, reducing it to NADH.

    • Net Reaction: Pyruvate + CoA + NAD+ →\rightarrow Acetyl-CoA + CO2CO_2 + NADH

  • Irreversibility: This reaction is highly irreversible with a very negative ΔG\Delta G, making it a key regulatory point.

  • Metabolic Boundary: This step represents a crucial transition from carbohydrate metabolism (pyruvate can be converted to glucose) to fatty acid metabolism (acetyl-CoA). Mammals cannot convert fat back into carbohydrates; thus, once pyruvate enters the acetyl-CoA pathway via this reaction, it's a "point of no return" for carbohydrate synthesis.

Regulation of Pyruvate Dehydrogenase

This enzyme is regulated by both product inhibition and phosphorylation/dephosphorylation:

  1. Competitive Inhibition by Products:

    • High concentrations of acetyl-CoA and NADH competitively inhibit the enzyme, signaling that subsequent processes are not consuming them.

  2. Phosphorylation (Inactivation):

    • Rising concentrations of acetyl-CoA and NADH activate a kinase, which phosphorylates the pyruvate dehydrogenase complex.

    • Phosphorylation occurs on serine, threonine, or tyrosine residues (primarily serine in this context), leading to enzyme inactivation.

    • This mechanism reinforces the shutdown of pyruvate conversion to acetyl-CoA when products accumulate.

  3. Dephosphorylation (Activation):

    • High pyruvate concentrations activate a phosphatase, which removes the phosphate group from the enzyme, leading to its activation.

    • Insulin, indicative of high blood glucose, also activates this phosphatase, promoting the conversion of glucose-derived pyruvate into acetyl-CoA for ATP production.

    • The overall logic is that high substrate (pyruvate, glucose via insulin) promotes the reaction, while high product (acetyl-CoA, NADH) inhibits it.

Citric Acid Cycle (Krebs Cycle / Tricarboxylic Acid Cycle)

  • Discovery: Discovered by Hans Krebs, who won the Nobel Prize with Lipmann in 1953. Krebs elucidated that this series of reactions forms a cyclic loop.

  • Overview: Acetyl-CoA (2C) combines with oxaloacetate (4C) to form citrate (6C). Through a series of reactions, two carbons are oxidized to CO<em>2CO<em>2, and electron carriers (NADH, QH</em>2QH</em>2) and GTP are produced, regenerating oxaloacetate to continue the cycle.

  • Key Outcomes:

    • The two carbons from acetyl-CoA are fully oxidized to CO2CO_2 and exhaled.

    • Coenzyme A is recycled.

    • Four electron pairs are captured by electron carriers:

      • 3 NADH (from 3 NAD+)

      • 1 QH2QH_2 (from 1 Q, Coenzyme Q, a hydrophobic molecule in the inner mitochondrial membrane)

    • 1 GTP (guanosine triphosphate) is produced, which is energetically equivalent to ATP.

  • Net Reaction (per acetyl-CoA):
    Acetyl-CoA + 3 NAD+ + Q + GDP + Pi →\rightarrow 2 CO<em>2CO<em>2 + 3 NADH + QH</em>2QH</em>2 + GTP + CoA

  • Directionality: The citric acid cycle operates in one direction (clockwise as typically drawn in diagrams). Humans lack the enzymes to catalyze the irreversible steps in reverse, meaning the entire cycle cannot run backward.

  • Oxygen Requirement: Oxygen is not directly consumed within the citric acid cycle itself. However, its presence is crucial for the cycle to continue because oxygen is the final electron acceptor in the electron transport chain (Topic 11), which regenerates NAD+ from NADH and Q from QH2QH_2. Without oxygen, these electron carriers would remain reduced, halting the citric acid cycle.

Regulation of the Citric Acid Cycle

Regulation ensures that the cycle's activity matches the cell's energy needs.

  1. Availability of Substrates: Increasing the concentration of key substrates like oxaloacetate and acetyl-CoA can increase flux through the pathway, particularly at the initial citrate synthase reaction.

  2. Competitive Inhibition by Products:

    • Products of the cycle, such as succinyl-CoA, competitively inhibit the enzymes that produce them (e.g., succinyl-CoA inhibits its own enzyme and also citrate synthase).

    • This is a general principle for metabolic pathways to prevent overproduction and maintain homeostasis.

  3. Allosteric Regulation:

    • Inhibitors: NADH is an allosteric inhibitor of several enzymes in the cycle, as it is a product. High NADH levels signal abundant reduced electron carriers, indicating that the cell has sufficient energy reserves and the cycle can slow down.

    • Activators: ADP is an allosteric activator. High ADP concentration implies low ATP levels, signaling an energy deficit. Activating the citric acid cycle increases the production of electron carriers and GTP, which can then be used to generate more ATP.

    • Calcium (Ca2+Ca^{2+}) can also act as an activator, signaling low ATP, though this is less emphasized.

    • The logic is paramount: if the cell needs energy (high ADP), the cycle is activated; if it has enough (high NADH), it is inhibited.

Intersections with Other Metabolic Pathways

  • The citric acid cycle and glycolysis are central metabolic pathways from which carbons can be diverted for synthesis or into which other molecules can feed.

  • Anabolism (Building Molecules): Intermediates from glycolysis and the citric acid cycle can be used to synthesize essential biological molecules:

    • Heme (used in hemoglobin, myoglobin)

    • Amino acids (some are non-essential and can be synthesized)

    • Nucleotides (components of DNA and RNA)

    • Cholesterol and other lipids

    • This means that glucose breakdown is not 100% efficient at producing ATP, as some carbons are used for biosynthesis.

  • Catabolism (Breaking Down Molecules): Other macromolecules can be broken down and enter these central pathways:

    • Amino Acids: Derived from protein breakdown, amino acids can feed into various points of glycolysis and the citric acid cycle.

    • This explains why proteins are considered an energy source (approximately 4 dietary calories per gram), similar to carbohydrates, as their breakdown products can be used to generate ATP.