Definition of Activation Energy: The energy required to convert reactants into products in a chemical reaction.
Relationship with Reaction Rate:
Higher activation energy means a slower reaction rate due to a steeper energy hill.
Lower activation energy results in a faster reaction rate.
Kinetic Stability of ATP
Thermodynamic Favorability vs. Kinetic Stability:
ATP hydrolysis to ADP and Pi is thermodynamically favorable but kinetically stable.
Meaning: Although ATP can break down to ADP, the reaction occurs slowly without intervention.
Enzymatic Acceleration:
Enzymes lower the activation energy barrier, allowing ATP hydrolysis to occur more rapidly.
Mechanism of Enzyme Action
Active Site Interactions:
Enzymes bind substrates at the active site, which requires energy to convert substrates into products.
The active site comprises amino acid residues that participate in weak non-covalent interactions (hydrophobic, ionic, hydrogen bonding).
Thermodynamic Energy Dynamics:
Binding of the substrate releases free energy, resulting in a negative change in Gibbs free energy (ΔG), effectively lowering the activation energy of the reaction.
Hexokinase and Glycolysis
Role of Enzymes in Glycolysis:
Hexokinase catalyzes the first reaction in glycolysis by transferring a phosphoryl group from ATP to glucose, forming glucose-6-phosphate (G6P).
Reaction Speed:
Without enzymes, ATP breakdown is slow. With hexokinase, it is expedited due to lower activation energy.
Relevance of the Reaction:
Breakdown of ATP to ADP during glycolysis occurs more quickly due to enzymatic action.
Thermodynamic and Kinetic Stability of ATP Hydrolysis
Thermodynamic Instability:
Breakdown of ATP is thermodynamically unstable, hence favorable.
Kinetic Stability:
High activation energy barrier (~+202 to +400 kJ/mol) makes the process kinetically stable and slow.
Use of Enzymes:
Enzymes efficiently catalyze ATP breakdown by lowering the energy barrier via binding energy interactions.
ATP Structure and Function
Phosphate Groups:
ATP consists of three negatively charged phosphate groups shielded by magnesium ions.
Nucleophilic Attack Mechanism:
Shielding allows the nucleophile to attack the gamma phosphate of ATP, facilitating the transfer to glucose.
Important for trapping glucose inside the cell since negatively charged molecules have no transporters to exit the cell freely.
Glucose Entry into Cells
Importance of Phosphorylation:
Conversion of glucose to G6P traps glucose inside cells and enables facilitated diffusion from high to low concentrations (blood to cell).
Glut Transporters:
Specialized transporters that allow glucose entry into cells, lined with polar residues for hydrogen bonding with glucose.
Isoenzymes of Hexokinase
Definition:
Isoenzymes (or isozymes) are different forms of enzymes with slight variations in amino acid sequences but perform the same chemical reactions in different tissues.
Comparison of Muscle Hexokinase vs. Liver Hexokinase:
Muscle hexokinase has a Km of 0.1 mM, allowing it to operate efficiently at normal blood glucose levels (4 mM).
Liver glucokinase, with a Km of 10 mM, has lower affinity and works best at high glucose concentrations following meals.
Enzyme Kinetics and Feedback Inhibition
Hexokinase Feedback Inhibition:
As glucose-6-phosphate levels rise, hexokinase is inhibited, demonstrating feedback inhibition.
Product inhibition occurs when high concentrations of the product regulate enzyme activity.
Mechanism of Hexokinase
Structure:
Composed of two lobes (large and small) with a cleft for ATP and glucose binding.
Induced Fit Model:
Enzyme undergoes conformational change to bring substrates closer together, facilitating the transfer of the phosphoryl group from ATP to glucose.
Conformational change also blocks water molecules from the active site to prevent premature hydrolysis of ATP.
Isomerization and Significance in Glycolysis
Step Two: Conversion of G6P to F6P (Fructose-6-Phosphate):
Phosphoglucose isomerase catalyzes conversion from an aldose to a ketose to allow cleavage.
Phosphorylation at the primary alcohol of carbon one is easier after conversion to a ketose structure.
Mechanism of Phosphoglucose Isomerase (PGI)
Initial Interaction with Substrates:
G6P binds and converts to linear form, enabling the aldehyde group to be exposed for catalytic activity.
Base catalysis by glutamate abstracts a proton to facilitate conversion to keto form, generating an enediol intermediate.
Acids and Bases in Active Site:
Glutamate acts in both base and acid catalysis during the conversion process.
Phosphofructokinase (PFK-1)
PFK-1 Role:
Catalyzes the committed step of glycolysis by converting F6P to F1,6BP (Fructose-1,6-Bisphosphate).
Characterized by having both regulatory and substrate-binding sites for ATP.
Energetics of Reaction:
Reaction is irreversible and has a large negative free energy, marking it as the rate-determining step in glycolysis.