TIM barrel Flashcards
What does “TIM barrel” stand for?
Triosephosphate isomerase barrel.
What is the general structure of a TIM barrel?
Eight α-helices and eight parallel β-strands that alternate along the peptide backbone, forming a closed barrel.
Which secondary structures alternate to form the TIM barrel?
β-strand → α-helix → β-strand → α-helix (repeated eight times).
What is the function of the β-strands in a TIM barrel?
They form the inner core (the barrel) of the structure, providing stability.
What is the function of the α-helices in a TIM barrel?
What is the function of the α-helices in a TIM barrel?
Why is the TIM barrel considered a versatile protein fold?
Because it can support a wide range of enzymatic functions — many different enzymes have evolved using this structure.
Where are the active sites typically located in TIM barrel enzymes?
At the C-terminal end of the β-strands, in loops connecting β-strands and α-helices.
What is the typical orientation of β-strands in a TIM barrel?
They are parallel β-strands (all running in the same N → C direction).
What reaction does triosephosphate isomerase (TIM) catalyse in glycolysis?
The interconversion of dihydroxyacetone phosphate (DHAP) into glyceraldehyde-3-phosphate (GAP).
Why is the DHAP → GAP conversion important in glycolysis?
Only GAP can proceed through the next steps of glycolysis to generate ATP. Converting DHAP ensures maximal energy extraction.
What type of reaction mechanism does TIM use to catalyse this conversion?
General acid-base catalysis — TIM stabilizes the enediol intermediate during the isomerisation.
Which key residues are involved in the active site of TIM?
Glutamate (acts as a base) and histidine (acts as an acid).
Describe the basic steps of the TIM-catalysed reaction.
- Glutamate deprotonates DHAP, forming an enediol intermediate.
- Proton transfer rearranges bonds.
- GAP is formed and released.
How does the TIM barrel structure support the enzyme’s catalytic function?
The β-barrel core stabilizes the protein.
The loops at the barrel top precisely position catalytic residues and bind DHAP tightly to prevent side reactions.
What is the importance of loop closure in TIM’s mechanism?
Upon substrate binding, a loop covers the active site (“loop closure”), preventing the enediol intermediate from decomposing — critical for catalytic efficiency and specificity.
Why is TIM often referred to as a “perfect enzyme”?
Its catalytic rate approaches the diffusion limit — meaning the reaction is as fast as substrate can reach the enzyme.
What is allosteric regulation?
Modulation of enzyme activity by binding an effector molecule at a site other than the active site, causing conformational changes.
Give an example of an allosterically regulated enzyme.
Phosphofructokinase-1 (PFK-1) in glycolysis, activated by AMP and inhibited by ATP.
What is feedback inhibition?
When the end product of a metabolic pathway inhibits an enzyme that acts earlier in the pathway.
What does turnover number (kcat) represent?
The number of substrate molecules converted to product per enzyme molecule per second under saturated conditions.
What is catalytic efficiency and how is it calculated?
Catalytic efficiency = kcat/Km; it measures how efficiently an enzyme converts substrate into product.
What is the effect of temperature on enzyme activity?
Increases reaction rate up to an optimal temperature; beyond that, enzyme denatures.
How does pH affect enzyme activity?
Each enzyme has an optimal pH; deviations can disrupt ionic bonds and denature the enzyme.
