L5-7: Enzymes and inhibitors Flashcards
What are enzymes?
Proteins
Catalysts
Have specific structures (stereo specific)
What are the most common type of drugs and poisons?
Enzyme inhibitors
What does Haemagglutinin do?
Facilitates entry of virus into host cells
How does Haemagglutinin enter the host cell?
Binds to sugars on the cell surface, changes conformation in host cell to fuse membranes and release virus in hosts
What is the enzyme that facilitates exit of the virus from the host cell?
Neuraminidase
How is the exit of the virus facilitated?
Binds to sugars on host cell surface and cleaves sialic acid sugars from cell glycoproteins, preventing virus sticking to cells
What are 2 anti-flu drugs?
Tamiflu and Relenza
How do the anti-flu drugs work?
They target the neuraminidase enzyme of influenza, drug has a similar shape so blocks active site
How does drug resistance arise?
From mutations (change in AAs in neuraminidase which allows infection of influenza)
How does Penicillin work?
Acts as a substrate which stops the crosslinking of peptides in glycan cell wall in bacteria (causes them to weaken and lyse)
How did bacteria become resistant to Penicillin?
Altered binding to beta-lactams so Penicillin is broken down with a beta lactamase enzyme
What is the key class of anti-viral drugs?
Nucleoside analogue drugs
What are examples of anti-viral drugs?
AZT- Azidothymidine (HIV)
Acyclovir (Herpes)
Remdesivir (COVID)
How do anti-viral drugs work?
Interact with RNA/DNA polymerases and reverse transcription enzymes (block active site and prevent further extension of DNA/RNA)
How do enzymes work?
Catalyse making and breaking of covalent bonds
Allow higher rate of reaction
Reactions can occur under milder conditions
Have high specificity
Regulate biochemical reactions
What is the equilibrium constant in a reaction, Keq, correlated to?
The energy release by the reaction (change in Gibbs)
What is the effect of catalysts on activation energy?
Lowers activation energy
What does free energy in a reaction determine?
Keq
How do enzymes impact equilibrium?
Speed up equilibrium but don’t change concentration
What is first formed before a product can form?
An enzyme-substrate complex
What are the properties of the mechanism of enzyme catalysis?
Proximity - close together
Orientation - correct relative rotation
Strain/distortion - binding puts strain on bond so easier for rection to occur
What is covalent catalysis?
Temporary covalent bond between enzyme and substrate
How do enzymes show specificity?
By having interactions between specific structural regions on the substrate and enzyme
What are the 2 different ways of an enzyme binding?
Lock and key- active site complementary to substrate
Induced fit- contact between active site and substrate changes shape allowing binding
What are the 7 different classes of enzymes?
Oxidorectuctases, transferases, hydrolases, lysases, isomerases, ligands, translocases
What is Kcat?
The number of substrate molecules converted to product by 1 enzyme in 1 second
When is an enzyme fastest in a reactions?
At the start
What does enzyme activity depend on?
Substrate concentration
What are the different effects of varying substrate concentration?
Low conc- number of substrate molecules determines how fast reaction takes place
Increased conc- more chance colliding (for catalysis to occur) & higher proportion of enzyme molecules will have bound to substrate
What happens when an enzyme is saturated?
The rate slows down as the increase of [S] does not make the reaction go faster
What happens in a first order reaction? (enzyme kinetics)
Rate of reaction increases proportionally with increased [S]
What is the equation derived from the relationship between velocity and substrate concentration?
𝑣= Vmax[S]/Km+[S]
Where 𝑣 is rate at a specified [S]
What is the Km constant?
[S] at half Vmax
What does Km mean?
It indicated the affinity of the enzyme for the substrate which is the stability of the ES complex
What do low and high Km mean in terms of affinity?
High Km= low affinity
Low Km= high affinity
What is the equation for Michaelis-Menten kinetics for a first order reaction?
V0=Vmax[S]/Km
What are the unis of Km?
Units of concentration (M)
What is Vmax and when does it occur?
It is the fastest rate at which an enzyme can work and only occurs at infinite [S]
How can Vmax be found?
Using Lineweaver-Burk plot
(slope = Km/Vmax)
What are the classes of enzyme inhibitors?
Irreversible (covalent complexes)
Reversable (useful + have an affinity)
-comp
-non-comp
-uncomp
Where do irreversible inhibitors usually bind?
To amino acid side chain or near active site
How do irreversible inhibitors work?
They bind permanently which inactivates the enzyme and prevents the substrate from binding
Name examples of irreversible inhibitors
Aspirin
Penicillin
Sulbactam
Allopurinol
AZT
Eflornithine
Sarin
5-fluorouracil
Exemestane
How does the irreversible inhibitor Sarin work?
It prevents the breakdown of NT acetylcholine as it binds to serine residue in acetylcholinesterase
How does Penicillin work?
Binds to Serine in penicillin binding proteins which irreversibly blocks the active site
What are the characteristics of a competitive inhibitor?
Competes with substrate for active site
Has similar structure to substrate
When bound blocks active site
How does competitive inhibition effect kinetics?
Increases Km
Vmax stays the same
What are examples of competitive inhibitors?
Tamiflu
Acarbose - type II diabetes (prevents digestion of natural carbs)
Statins- target hepatocytes and inhibit HMG-CoA reductase
What are the characteristics of non-competitive inhibition?
Binds away from active site
Modifies reaction rate
Binds substrate with same affinity
How does non-competitive inhibition effect kinetics?
Km unaltered
Vmax decreased
What is uncompetitive inhibition?
Occurs with multi-substrates
Binds only to ES complex
How does uncompetitive inhibition effect kinetics?
Km decreases
Vmax decreases
(ratio stays the same)
What reaction conditions influence enzyme activity?
Temperature
pH
Salts
Other chemicals
Molecular crowding
How does pH change enzyme activity?
3D structure changes
Groups involving in binding substrate or catalysis change charge
How does temperature change enzyme activity?
Change diffusion rate
Flexibility of active site
Stability of enzyme (denaturation)
What is molecular crowding and how does it impact enzyme activity?
Large proteins diffuse slower than small substrates/products
Cells can increase amounts of slower enzyme
Pathways with volatile intermediates cell can trap them
What is the carboxysome?
Carbon concentration mechanism
Balances activity of carbonic anhydrase and rubisco
Protein shell to localise CA next to rubisco
Traps CO2
