EnYems And Flashcards
Why must metabolism be accelerated by enzymes?
To operate on a time scale relevant to life
What does thermodynamics NOT a tell you
Speed of a reaction
What does he say (3 things) about this reaction:
ATP ADP + Pi
- Thermodynamically favourable so it’s spontaneous (-🔺G)
- Incredibly slow in absence of catalyst
- However in the context of ATP this is also a good thing because we don’t want to deplete energy store really quickly
How much does Triose Phosphate Isomerase (TPI), an enzyme in glycolysis speed up the reaction
- what’s the rate of the uncatalysed reaction
- catalysed reaction
- rate enhancement
Uncatalysed: one product/day
Catalysed: 4300 products/second
Rate enhancement: 1000 million fold
Slowest uncatalysed reaction
Synthesis of UMP (a pyramiding nucleotide)
1 reaction per 45 million years
When the synthesis of UMP is catalysed
Catalysed
Reaction enhancement
Catalysed - 39 products/ second
Rate enhancement: 1.4x10^17 fold
What is the synthesis of UMP catalysed by
OMP decarboxylase (decarboxylates orotidine monophosphate to UMP)
Why are uncatalysed reactions so slow
Because they go from a substrate to product via an intermediary state (the transition state)
Transition state
Transitory molecular structure that is no longer the substrate but not yet the product
What’s the least stable species in a reaction?
The transition state
What has the highest free energy in a reaction?
The transition state
What will free energy be like in a spontaneous reaction
Free energy of products will be lower than free energy of reactants
What does an enzyme do in terms of free energy?
It reduces the free energy of the transition state
How do enzymes accelerate a reaction in terms of transition state and activation energy?
They stabilise the transition state , reducing the activation energy required to reach it
Enzymes affect the ___ NOT the ___
Enzymes affect the KINETICS not the THERMODYNAMICS
Delta G remains the same
Two unaltered things when an enzymes acts
- 🔺G
2. Equilibrium position
What does an enzyme do in terms of equilibrium
Accelerates how long it takes to reach equilibrium
Simplistic analogy of how enzymes stabilise transition state
Enzyme complementary to transition state
But leaving enough room so that once the conformational change occurs the substrate is completely complementary to the enzyme (magnets arranged so they anticipate the shape of the broken sticks)
Because then by being completely complementary and snug with the transition state it makes the transition state more stable which reduces free energy required to reach it
What part of an enzyme shares similar features in all enzymes
The active site
General features of the active site
- Crevice in the enzyme structure
- AA side chain catalytic groups
- Water excluded
- Hydrophilic side chains
What are catalytic groups on enzymes
Within active site key AA side chains known as catalytic groups are precisely orientated and around the bound substrate
What is meant by water is excluded in the active site of an enzyme
Crevice is mainly lined with hydrophobic amino acids so water excluded unless it’s a reactant
What is meant by there are SOME amino acids with hydrophilic side chains in the enzyme active site?
There are a few and these polar residues acquire special properties essential for substrate binding or catalysis
What is meant by some hydrophilic residues in active site of enzyme
Some but they acquire special properties essential for substrate binding or catalysis
What’s the driving force behind protein folding
Hydrophobic residues make protein fold to have a hydrophobic core
Why are proteins preferred as a catalyst rather than RNA
proteins can fold specifically due to hydrophobic action of residues
Proteins can be made of 20 different amino acids which all provide improved chemistry for a catalyst
Why are proteins better catalysts than RNA
Proteins can fold because of natural hydrophobic action
Improved chemistry set of 20 AA side chains
Covalent catalysis
Breaking or making of covalent bonds between enzyme and substrate
(Typically active site has a highly reactive group that become temporarily covalently attached to the substrate)
Acid base catalysis
A molecule other than water acts as an acid (proton donor) or a base (proton acceptor)
In the context of enzymes metal ions are usually
Positively charged
How can metal ions assist catalysis (3)
- Help bind substrate
- Stabilise negative charges on reaction intermediates
- Increase acidity of interacting molecules (promotes the molecules role in acid base catalysis)
Catalysis by enhanced proximity of reactants
Bring two molecules close together can enhance transfer of function groups
4 mechanisms enzymes commonly employ to catalyse specific reactions
- Covalent catalysis
- General acid base catalysis
- Metal ions catalysis
- Enhanced proximity
Acid-base catalysis example
How does chymotrypsin catalyse the hydrolysis of peptide bonds with this mechanism?
Chymotrypsin attacks the unreactive carbonyl in the peptide bond with a nucleophile.
And forms a covalent bond
What is the nucleophile in chymotrypsin that aids catalysis the peptide bond
Serine 195 residue
What’s the catalytic triad (chymotrypsin action)
Asp 102
His 57
Set 195
How does the serine 195 in chymotrypsin become a nucleophile?
+ how is the catalytic triad involved
His attracts H in serine OH group
Serine now a nucleophile because it’s lost the H from its OH group so is just the O- (forms a charged alkoxide ion)
Asp forms H bond with His to allow His to more easily accept to H from the Ser
Then sweetlove illustrates a catalytic mechanism using
Chymotrypsin as an example
Looks at how it uses covalent and acid base catalysis
Example of base catalysis in chymotrypsin action
Chymotrypsin uses histidine residue as a base catalysis to increase nucleophilicity of Seine
What can the kinetics of many enzyme catalyse reactions be mathematically described by?
Michaelis-Menten model
Why is it useful to know Vmax and Km
If u know what Vmax and Km are then u get define mathematically what the enzyme will do for any given substrate concentration
How to get Vmax and Km
Use purified enzyme and do experiments
When [S] and Km are the same what is the velocity ?
Half of the V max
What is Km a measure of
Affinity at which enzyme binds substrate
Lose Km means what
Tighter it’s binding it’s substrate and so faster reaction will go go a given substrate concentration
Why are enzyme kinetics important?
Allows us to make predictions about how enzyme systems behave
Sumamry of the confusing bit!
- Enzymes control rate of reactions
- If rate of one recation is faster than another so that more product is used than made by the supplying reaction then it wouldn’t wortk
- So enzymes keep all reactions at a steady state
- Enzyme kinetics have evolved so that metabolite concentrations are all at a steady state
- This is important because it keeps metabolite concnetrations in a low range otherwise the cell would become staurated and there wouldn’t be enough water to sufficiently hydrate everything
Example of enzyme kinetics being an important tool for biotechnology
involved in biosynthesis of lysine (animal food addidative). If they’ve got all these constants then they can try and optimise the system.
EG. Try and optimise Kms of enzymes to engineer different affinity of substrate and see how it would affect production of lysine
In a system of enzymes the kinetic response of enzymes to changes in substrate concentration means …
the system tends towards a steady state
Vmax and Km are selected by evolution because
need to keep metabolite concentrations at steady state relatively low so solvency capacity of cell is not exhausted
