The Behavior of Proteins:Enzyme Kinetics I
Chapter 6 - pgs. 143-151
Outline
• enzymes kinetics (speed/rate) and thermodynamics (making the universe happy) • determining the rate of a catalyzed reaction
• enzyme-substrate binding
• CATALYSIS is the act of _____________________ up a chemical reaction • when my 3.5-year old takes 15 minutes to put on her shoes…
- I become the catalyst by saying “c’mon now! hurry it up!” • without catalysis, the reaction would occur much slower
• but – KEY CONCEPT – catalysis does not __________ the _______ _________________! - the same thing happens with or without catalysis
- catalysis just makes it happen faster • those shoes will get put on either way…
• of all the things proteins do – and they do plenty – catalysis is the most important
• without protein catalysts – ENZYMES – chemistry would take too long for life to be possible • enzymes are globular proteins
- but it’s important to note that not all proteins are enzymes - and not even all globular proteins are enzymes
• it’s also important to mention that specific types of RNA molecules (called ______________) can achieve catalysis
- these are not proteins; they’re RNA
- but _____________________ are beyond the scope of this course • enzymes are amazing catalysts
- they speed up chemical reactions on levels we can’t even conceive - enzymes are the best catalysts out there
- nothing speeds up chemistry better…
• enzymes also offer an amazingly high degree of __________________________
Kinetics vs. Thermodynamics
• these are two different and distinct properties, but closely related - and this is true of all reactions (catalyzed or uncatalyzed)
• let’s first consider thermodynamics
• the universe likes free energy (like __________)
- free energy, when released, makes things move more, move faster, be more chaotic, be more random
- and if the universe loves one thing… it’s randomness (_________________)
• so the universe likes and encourages things that _________________ free energy
• things that absorb free energy (trap it, bottle it up) are ‘stealing’ that free energy from the universe
- as a consequence, things move a little less, a little slower - this makes the universe sad….
• if we want to determine whether or not the universe will like a given chemical reaction, we first
need to determine if that reaction ____________________ free energy
• to do that, we need to look at the free energy contained in the reactants (what goes into the chemical reaction) and the free energy contained in the products (what comes out of the reaction)
• we call the _________________ in free energy ΔG - Δ for change or difference
- G for heat or free energy (because scientists are IDIOTS…)
• and so ΔG = (free energy of the products)–(free energy of the reactants)
• if the free energy of the products is lower than the free energy of the reactants ΔG will be negative (-) and free energy will be released
- does the universe like it when this happens…? • which of the reactions below does the universe like…?
• reactions that have a –ΔG are called ___________________________ reactions - the universe wants this reaction to happen because it will release free energy • the reaction of glucose and oxygen to make carbon dioxide and water is enormously
__________________ and spontaneous (ΔG = -2880 kJ/mol) - this is how we get our energy
- why isn’t anything happening…? my eyebrows should be gone! • when we consider ΔG, all we are looking at is this…
• we are not looking at the ____________ between the two
• for this reaction to go from reactants to products, there’s a pretty big hill to climb – an
__________________ hill
• the universe may want this reaction to occur – because when it’s all done free energy will be released
- but to get there, the universe must first pump quite a bit of energy into the reaction
- it needs to _______________ energy in this reaction - and the universe is not much into investing energy
- SPONTANEOUS does not mean ________________________________!! • it may be ‘thermodynamically favorable’ to get from reactants to products
- but not if I put a mountain of free energy between them
• this mountain of free energy is called the ENERGY OF ______________________ and abbreviated ΔG‡
• enzymes DO NOT affect ΔG! - ΔG is path independent!!!
• enzymes lower the energy of activation making that hill _______________
- all catalysts improve the chances a chemical reaction will occur simply by lowering that ΔG‡ • and so, an enzyme can sped up how fast a reaction occurs (kinetics) by making it easier to
occur
• but, an enzyme does not change the energy of the reaction (thermodynamics) by affecting the ENERGY OF ACTIVATION
- you were going from the Yaletown to the Downtown hotel, mountain for not - same products go to the same reactants, with or without the enzyme
• enzymes do not affect ΔG, only ΔG‡
The Transition State • why…?
- catalyzed or not, why does this energy (ΔG‡)need to be pumped into the reaction? • chemical reactions turn reactants into product
• the reality of this is reactant must be pulled, stretched, pushed, squeezed, hammered, ripped, and pummeled into product
- this takes energy
• at the peak of the graph (ΔG‡) lies our TRANSITION STATE
• the transition state is a ____________ structure that is no longer reactants, but also not quite product
- it is something different on its way to becoming product - remember the bread machine analogy…?
- water, flour, salt, eggs are the reactants - freshly baked bread is the product
- ____________ is the transition state - it certainly is NOT reactants
- but also not quite product
- it’s something inbetween that is very much on its way to becoming product… • so, then, let’s define the ENERGY OF ACTIVATION a bit better:
- ΔG‡ is the amount of free energy needed to reach the transition state (once you get there, it’s all downhill – literally!)
Enzyme Kinetics
• kinetics refer to the speed or rate of a chemical reaction
- we usually determine this rate and express it in terms of a change in the concentration of reactants or product in a given interval of time
- reactants should disappear and product appear as the reaction proceeds A + B → P
• rate = -Δ[A]/Δt = -Δ[B]/Δt = Δ[P]/Δt
- remember: Δ[A] and Δ[B] are negative already, so the negative of a negative gives a positive rate
• we’ll skip the actual RATE EQUATION because it adds little to the understanding and lots to the confusion
- rate = k[A]f[B]g
• the ___________________ ORDER of a chemical reaction refers to the number of molecules involved in the rate of the reaction
- what the rate of the reaction depends on
A → P
• only A is involved in the rate – A becomes P
- this is a ____________ order reaction overall (only __________ molecule involved) - this is a first order reaction with respect to A (from A’s perspective, only one molecule is
involved – itself…)
• an example of this is the decay of radioactive 32P 32P → decay products
- __________ order reaction
- the only thing the rate of this reaction depends on is 32P
A + B → C + D
• this is a _____________ order reaction overall - A and B are both involved in the chemistry • this is a first order reaction with respect to A • this is a first order reaction with respect to B
- from each of their perspectives only one of them is needed for the rate of the reaction Glycogenn + Pi → Glucose-1-phosphate + Glycogenn-1
• ____________ order overall – both glycogen and orthophosphate are involved in the chemistry
• first order from the perspective of each reactant – only one of each reactant is needed for one of the other
• what is a ____________ order reaction?
- when the rate of the reaction is completely ___________________ of the concentration of the reactant(s)
• enzymatic reactions can be _______ order when the concentrations of the reactants (substrates) are so high that they saturate all the enzymes
- in this case the reaction is proceeding as fast as possible - the rate is as high as it can possibly go
- and so the rate of the reaction is independent of anything else
• cars waiting at a six-lane toll plaza to cross a two-lane bridge is a zero order event
- the rate of cars crossing is constant and will not change if some cars are added or removed
Enzyme-Substrate Binding
• if enzymes are going to work on reactants to make them products, then enzymes must first bind reactants – SUBSTRATE(S)
- this results in the enzyme-substrate complex
• in this complex, the enzyme pulls, stretch, rips, hammers, and squeezes the substrate into the transition state
- once the transition state forms, it will collapse into product all on its own (because this releases tons of energy)
E + S → ES → E + P
• the substrate usually binds to a small _______________ in the enzyme called the ACTIVE SITE - this cleft/crevice holds the proper substrate perfectly and does not fit well with anything else –
____________________________
• the substrate makes numerous contacts with the _____________________ and ____________ components of the enzyme’s active site
- these numerous contacts give the specificity
- these numerous contacts allow the enzyme to pull the substrate to the transition state • for a long time, the LOCK-AND-KEY model was used to explain this
- but this model has since been proven to be inaccurate • the newer model – INDUCED FIT – is more correct
- the active site is quite __________________
- substrate binding induces conformational changes that allow the active site to _______ around the substrate
• the induced fit model predicts something very important
- it’s good for an enzyme to _________________ perfectly with its substrate - the lock-and-key model predicts a perfect fit
- why is it good for the fit to not be too perfect…?
• the ES complex forming must be thermodynamically favorable - otherwise, it would never happen
• but, if it were too favorable, the ES complex would never move on to catalysis
• if this fit were perfect perfect perfect – enzyme and substrate would meet, bind, and never let go happily ever after
• in fact, enzymes usually bind best to the ____________________ _________________
- the enzyme likes binding to the _________________ __________________ so much, it will pull, stretch, squash and pummel substrate into it…
• and so, the enzyme binds substrate
- the enzyme lowers the energy of activation by working hard to make the transition state form - once the transition state forms, it will rapidly collapse into product
- the transition state is a high energy molecule (high maintenance) - the universe hates high maintenance molecules
- and so, the transition state exists extremely briefly
- once product is formed, it fits so poorly into the enzymes active site that it is quickly released - the enzyme is now ready to start the whole process over again with another substrate
molecule
- KEY CONCEPT: enzymes ___________________ – over and over and over again • different enzymes achieve catalysis in different chemical ways
• but they all speed up the rate of the reaction by lowering the ΔG‡
Summary
• CATALYSIS is the act of speeding up a chemical reaction
• the universe likes and encourages things that _________________ free energy • if the free energy of the products is lower than the free energy of the
reactants ΔG will be negative (-) and free energy will be released
- this is a ______________________ reaction (but not necessarily a fast one!) • the energy of activation is the mountain of free energy that must be
- all enzymes do is _______________ the energy of activation - they change nothing else about the chemical reaction
• we did some dry kinetics (zero order, first order & second order rxns) • the lock-and-key model vs. the induced fit model
