Enzymes Flashcards
Describe the mechanism of action of enzyme molecules
-specificity:
The active site of an enzyme is a specific shape, depending on the reaction that it catalyses, meaning that other molecules won’t fit into the active site
-active site:
The area on an enzyme to which the substrate binds
-lock and key hypothesis:
The theory of enzyme action in which the enzyme active site is complementary to the substrate molecule, like a lock and key. The enzyme is held in one place so the reaction can happen.
-induced-fit hypothesis:
The theory of enzyme action in which the enzyme molecule changes shape to fit the substrate molecule more closely as it binds to it. The substrate is held by the enzyme’s oppositely charged R-groups. It lowers the activation energy by weakening bonds in substrate when the enzyme changes shape.Products are a different shape, don’t fit, so move away from active site.
-enzyme-substrate complex:
The intermediary formed when a substrate molecule binds to an enzyme molecule
-enzyme-product complex
The intermediate structure in which product molecules are bound to an enzyme molecule
-lowering of activation energy:
Enzymes reduce the activation energy (by destabilising the bonds in the substrate) so the reaction can proceed at a much lower temperature
Effects of pH on enzyme activity
Narrow pH range
Low pH = lots of H+ ions
H+ ions have a positive charge
Either extreme of H+ ion concentration can interfere with the hydrogen and ionic bonds holding the tertiary structure together.
The pH affects the charge of the amino acids at the active site, so the properties of the active site change and the substrate can no longer bind
At high pH values, a –COOH group will dissociate to become a charged –COO- group
Effects of temperature on enzyme activity
Up to a certain point, increasing temperature will increase the rate of reaction, as higher Ek means there will be more collisions between enzymes and the substrate, and more of these collisions will have the required activation enthalpy for the reaction to proceed.
But heat makes enzyme molecules vibrate, putting strain on the inter-molecular bonds, weaker bonds (hydrogen bond and ionic bonds) may break.
In enzymes there are many hydrogen and ionic bonds holding the tertiary structure, and especially the active site, in place.
As the heat increases, more bonds are broken
The tertiary structure disintegrates further
The rate of reaction decreases
If enough of these bonds are broken, the entire tertiary structure will unravel and the enzyme will stop working
This is not reversible and is known as denaturation
Effect of enzyme concentration on the rate of an enzyme controlled reaction
As enzyme concentration increases, the rate of reaction increases linearly as there are more active sites are available, until the substrate concentration becomes a limiting factor and the rate stops increasing
The effect of substrate concentration on the rate of an enzyme controlled reaction
As the substrate concentration rises, the rate of reaction rises because there are more substrate molecules to react. At higher concentrations, all of the active sites become filled, so the rate of reaction remains the same and the enzyme concentration becomes a limiting factor
Experiment for effects of pH on enzyme activity
Amylase catalyses starch to maltose
starch-agar plate (mixing starch with agar) left to set in a Petri dish.
Cut holes into the plate using a cork borer.
Into each hole place the same volume of a different pH buffer solution
Add an identical volume of stock amylase solution
One hole with distilled water as a control
Incubate for 24h in a dry oven at 35 ̊C
Flood the plate with an iodine solution (shows u catalyses starch as blue) and rise with water
Measure the diameter of the cleared zone- this gives an indication of how much substrate has been turned into product
Repeat at least three times, identify anomalous results, find a mean, compare results
Experiment to investigate the effect of temperature on enzyme activity
Catalase catalyses hydrogen peroxide to water and oxygen gas.
samples of potato tissue (contains catalase) using a cork borer, cut discs of equal thickness
Place an equal number of discs in several test tubes each tube is put in a WATER BATH (range 20-80 ̊C)
equal volume of pH 7 buffer and hydrogen peroxide into other test tubes in each water bath. Allow to equilibrate.
Taking each potato test tube in turn, add peroxide/buffer mixture to them, then fix a stopper and gas syringe
Time how long it takes for the gas syringe to fill the same volume
Calculate the rate by rate=1/time
Repeats!
Experiment to investigate effect of enzyme concentration on enzyme activity
Samples of potato tissue using cork borer, slice into discs of equal thickness
Place a different number of DISCS into several test tubes, put in a water bath at one temperature
pH buffer equal volumes and hydrogen peroxide solution equal volumes into separate test tubes, into same water bath- allow to equilibrate.
Add hydrogen peroxide and pH buffer to the potato disc tubes, fix a stopper and a gas syringe.
Time how long it takes to reach the same volume of gas produced.
Calculate rate=1/time
Experiment to investigate substrate concentration on enzyme activity
Samples of potato tissue using cork borer, slice into discs of equal thickness
Place the same number of discs into several test tubes, put in a water bath at one temperature
pH buffer equal volumes and HYDROGEN PEROXIDE SOLUTION different volumes into separate test tubes, into same water bath- allow to equilibrate.
Add hydrogen peroxide and pH buffer to the potato disc tubes, fix a stopper and a gas syringe.
Time how long it takes to reach the same volume of gas produced.
Calculate rate=1/time
Effects of competitive inhibitors on the rate of enzyme-controlled reactions
similar shape to substrate molecule
they occupy the active site, forming enzyme-inhibitor complexes. complexes do not lead to the formation of products-inhibitor is not identical to the substrate.
level of inhibition depends on concentrations of inhibitor and substrate.
substrate molecule increased, level of inhibition decreases (a substrate molecule is more likely than an inhibitor molecule to collide with active site).
Most competitive inhibitors don’t bind permamently to the active site. short period of time and then leave
action is reversible, as the removal of the inhibitor form the reaction mixture leaves the enzyme molecule unaffected.
Effects of non-competitive inhibitors on rate of enzyme-controlled reactions
Don’t compete with substrate molecules for a place in the active site. attach to the enzyme away from active site
attachment of non competitive inhibitors distorts the tertiary structure of the enzyme molecule, shape of the active site changes. Substrate no longer fits into active site so the enzyme-substrate complexes cannot form- reaction rate decreases.
level of inhibition depends on the number of inhibitor molecules present. Enough inhibitor molecules to bind to all of the enzyme molecules presend, then the enzyme controlled reaction will stop. Changing the substrate concentration will have no effect on this form of inhibition, as they don’t compete for same space.
Most non-competitive inhibitors bind permanently to the enzyme molecule. The inhibition is irreversible, and any enzyme molecule bound by inhibitor molecules are effectively denatured.
Importance of cofactors and coenzymes in enzyme-controlled reactions
Cofactors
Ions that increase the rate of enzyme-controlled reactions. Their presence allows enzyme- substrate complexes to form more easily.
They include coenzymes, prosthetic groups and inorganic ion cofactors.
Coenzymes
Small, organic, non-protein molecules
bind for a short period of time to the active site.
bind just before, or at the same time as the substrate binds.
coenzymes can take part in the reaction, and are changed in some way.
Unlike the substrate, coenzymes are recycled back to take part in the reaction again.
often carry chemical groups between enzymes so they link together enzyme-controlled reactions that need to take place in sequence.
Some coenzymes are permanent parts of the enzymes- prosthetic groups. These contribute to the 3D shape and charges of the enzyme.
Metabolic poisons can be described as…
And describe the action of one named poison
Enzyme inhibitors
Potassium Cyanide acts as a non-competitive inhibitor of the enzyme cytochrome oxidase, which is involved in the oxidation of ATP. When this is inhibited, aerobic respiration cannot occur, and so the organism can only respire anaerobically, which leads to a build up of lactic acid, toxic to the cells.
One application of enzyme inhibitors
Some medicinal drugs work by inhibiting the activity of enzymes
What is an enzyme?
A globular protein
With a specific tertiary structure
Which catalyses metabolic reactions in living organisms
And can be infra cellular or extracellular
