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What is an active site
Indented area on the surface of an enzyme molecule. It's sites shape is complementary to the shape of the substrate molecule.
What is a catalyst?
Chemical that speeds up the rate of a reaction without being used up and is reusable at the end of the reaction.
What are enzymes?
They are biological catalysts as they speed up the rate of metabolic reactions in living organisms.
What is the number of reactions an enzyme molecule can catalyse per second known as?
Turn over number.
Why are enzymes so remarkable compared to chemical catalysts? (Think chemistry GCSE!)
Chemical catalysts usually need ;
- High temperatures (enzymes do not)
- Increased pressures (enzymes do not)
- Extreme pH's (on average enzymes work near 7.0)
- Enzymes are more specific than chemical catalysts (enzymes rarely make mistakes and do not produce unwanted by products)
Why is the active site important?
- Tertiary structure of active site is complementary to the shape of the substrate molecule(s), allowing the catalysing of a substrate to occur
- Therefore each enzyme is highly specific
What can change the shape of an enzyme and its active site?
- pH
- Temperature
Why is the enzymes active site changed by these factors?
The bonds holding the proteins in their tertiary structure could be broken.
What types of environment can enzymes work in?
Intracellularly or extracellularly.
When enzymes are secreted extracellularly in a permeable environment, can those enzymes undergo diffusion?
Yes
What are the two different metabolisms enzymes can work with?
- Catabolic metabolic pathways
- Anabolic metabolic pathways
What is a catabolic pathway?
Substrates are broken down into smaller molecules, releasing energy
Graph of energy changes of catabolic reactions:
What is an anabolic pathway?
Substrates are joined to synthesise larger molecules. Energy is needed
Graph of energy changes of anabolic reactions:
What are the reactants, intermediates and products known as?
Metabolites
What are the reactants and intermediates known as for enzymes?
Substrates
Where is catalase found?
Nearly all living organisms exposed to oxygen.
Why is catalase needed?
Protects cells from damage by reactive oxygen.
What is the reaction pathway of catalase?
Hydrogen peroxide --> oxygen + water
Structure of catalase? Also is it the Slowest/average/fastest enzyme?
- Four polypeptide chains, containing a haem group with iron
- Fastest acting enzyme, with a turnover rate of 6 million per second
Where is catalase found in eukaryotic cells?
Inside small vesicles known as peroxisomes
What cell that is found in the bloodstream uses catalase in the human body?
White blood cells; to help kill invading microbes
What is the optimum pH and temperature of catalase for humans?
- pH 7.0
- Temperature; 45 degrees Celsius
What are the two enzymes examples that are produced for extracellular purposes in the human body?
- Trypsin
- Amylase
(I know there are a lot more we all need to know for that uni degree, but you also need to know how to win the game against the A level spec)
Where is the secretion of trypsin?
Produced in the pancreas, acts in the lumen of the small intestine.
What is the function of trypsin?
Digests proteins into smaller polypeptides through hydrolysis of peptide bonds.
Optimum pH of trypsin?
Between 7.5 and 8.5 pH
Where is the secretion of amylase?
Produced in the salivary glands, acting in the mouth.
Produced in the pancreas, acting in the lumen of the small intestine.
Function of amylase?
Digests polysaccharide starch to disaccharide maltose.
What is a cofactor?
A substance that needs to be present to ensure an enzyme catalysed reaction takes place at an appropriate rate.
What are three examples of cofactors?
- Prosthetic groups
- Mineral ion cofactors
- Organic coenzymes
Which cofactor(s) is permanently part of the enzyme structure?
Prosthetic groups
Which cofactor(s) is temporarily part of the enzymes?
- mineral ion cofactors
- Organic coenzymes
What is a complex formed by temporary binding of an enzyme and substrate molecule(s) during enzyme and substrate reaction called?
Enzyme-substrate complex
Why do some enzymes require cofactors?
As some enzymes requires a small non-protein molecule to work;
- To supply energy
- Ensure more efficient binding of the substrate and enzyme
Example of an enzyme that uses a zinc ion as a prosthetic group?
Carbonic anhydrase.
Reaction of carbonic anhydrase?
H2CO3 <--> CO2 + H2O
Why is carbonic anhydrase required?
Helps carbon dioxide to be carried out in blood from respiring tissues to the lungs.
How can certain ions temporarily bound to enzymes or substrate (mineral ion cofactors, coenzymes) increase the turnover rate?
Cofactors ease the formation of enzyme-substrate complexes
What can some co-factors do as part of their function?
- Bind to substrates to enable a correct shape for the enzyme they bind to. Known as Co-substrates
- Some co-factors change the charge distribution on the ;
* surface of the substrate molecule
* surface of the enzyme's active site
Therefore the temporary bonds in enzyme-substrate complex are easier to form.
What ions does amylase need to function?
Chloride ions
What can metallic ions be as cofactors?
- Permanent cofactors forming prosthetic groups
- Temporary cofactors to aid formation of enzyme substrate complexes
Do all cofactor ions need to be metallic?
No, some are non-metallic cofactor ions
What are coenzymes?
Small organic non-protein molecules that bind temporarily to the active site of enzyme molecules, sometimes to supply energy or a molecule.
Do coenzymes become chemically changed after a reaction?
Yes, they also need to be recycled to their original state by a different enzyme.
What coenzyme is B12 derived from? Disease without it?
cobalamin. Pernicious anaemia
What coenzyme is Folic acid derived from? Disease without it?
Tetrahydrofolate. Megablastic anaemia
What coenzyme is Nicotinamide derived from? Disease without it?
NAD, NADP. Pellagra
What coenzyme is Pantothenate derived from? Disease without it?
Coenzyme A. Elevated blood pressure triglyceride levels.
What coenzyme is Thiamine derived from? Disease without it?
Thiamine pyrophosphate. Beriberi
What is an enzyme-product complex?
Enzyme molecules with product molecule(s) in its active site. Joined by non-covalent forces
What is an enzyme-substrate complex?
Enzyme molecule with substrate molecules in its active site. Joined by non-covalent forces
Two different models/hypothesis' of how enzyme-substrate complex forms?
1. Lock and key hypothesis
2. Induced fit hypothesis
How does the lock and key hypothesis work?
1. Substrate molecules and enzyme molecule each have kinetic energy and are constantly moving randomly
2. Substrate molecules collide with an enzyme molecule
3. Substrate(s) binds to it's complementary enzyme via the enzymes active site.
4. Temporary hydrogen bonds hold the two molecules together. Enzyme-substrate complex is formed.
5. Product(s) molecules are released as they leave the active site.
What is the lock and what is the key in the lock and key hypothesis?
Lock - Enzymes active site
Key - substrate(s)
Disadvantage of the lock and key hypothesis?
Doesn't explain how the transition state, the ES complex, is stabilised
What is different about the induced-fit model compared to the lock and key model?
Enzyme does not have a rigid structure. Substrate induces the active site.
How does the induced fit model/hypothesis work?
1. Substrate fits into the enzymes complementary active site, the active shape moulds around the substrate molecule
2. As an Enzyme substrate molecule starts to form, the R-groups shape changes subtly to exactly fit the substrate
3. Moulding enables substrate to bind more effectively
4. Enzyme-substrate formed as non-covalent forces (i.e. hydrogen bonds, van der waal, polar attractions) occur
5. Substrate molecules converted to product molecule(s) and enzyme-product complex is formed
6. Product(s) has a slightly different shape to substrate molecules, detachment occurs and product(s) released
7. Enzyme can convert another substrate(s)
Can digestion be used synonymously with hydrolysis?
Yes.
But for the sake of you getting 100%, know when to say each, as sometimes in an exam you will only get the terminology mark for hydrolysis.
What does digestion actually mean?
To break down
What is the usual way for an activation energy to be met in enzymes? What does the form of energy do?
Thermal energy, as it provides the kinetic energy
Why do enzymes actually need the kinetic energy?
To have more frequent successful collisions (due to activation energy being met) and therefore more enzyme-substrate formations can happen (more reactions)
What happens if the temperature is raised too much in a solution of enzymes and its substrate.
Enzymes will denature and any lipids will melt.
What is the main feature of an enzyme? (why do we need catalysts).
They lower the activation energy. Therefore excessive heat is not needed.
What is Q10?
Temperature coefficient; calculated through
Q10 = dividing the rate of reaction at (Temperature + 10) celsius/rate of reaction at (temperature) celsius
What happens if a substance is heated?
- Extra thermal energy causes the molecules to move faster
- Increases the rate of collisions between molecules
- Increases the force with which they collide at
What happens if a solution containing enzymes and substrates increases in temperature? (before the optimum temperature)
- Both types of molecules will gain kinetic energy and both will move faster
- Increase number of successful collisions
- Rate of formation of Enzyme-Substrate complexes increases
- Rate of reaction increases as number of enzyme-product complexes increases and therefore the turnover rate increases.
- At a specific temperature, enzyme's rate of reaction will meet its maximum. Known as the optimum temperature
Do all collisions between enzymes and substrates occur in a Enzyme-substrate complex?
No! Only collisions that are "successful" where both substrate and enzyme are in the correct orientation and enough thermal energy to overcome the activation energy will occur in an Enzyme substrate being formed.
What happens to an enzyme when temperature goes over the optimum level?
1) Hydrogen and ionic bonds in the tertiary structure of the active site may get broken down
2) Active site begins to change; substrate molecules may not fit so well into the active site, rate of reaction decreases
3) As heat increases (as is temperature), shape of enzyme's active site irreversibly changes. Enzyme is no longer complementary to the substrate.
4) Reaction cannot proceed at all
5) All enzymes are now denatured.
When the enzyme has heat applied to it over the optimum rate does the peptide bonds break?
No, the peptide bonds do not break while hydrogen bonds do
What does this mean to the enzymes primary structure?
Primary structure is still intact though tertiary structure has dramatically changed.
Is the enzyme still folded after denaturation?
No, the enzyme has unfolded, tertiary structure changed completely
Do all enzymes have to work at good temperatures? (i.e. between 20 and 40 celsius)
No, some enzymes can work at low temperatures and some are heat stable and work at very high temperatures
How can some enzymes be adapted to being more heat stable?
They possess more disulphide bridges
What are the two rate of reactions equations?
Rate of reaction (seconds) = 1/time taken to reach an end point
Rate of reaction (dm^3/s) = (Product formed or substrate consumed)/time
What is Q10 usually for chemical reactions?
2
As this is the usual Q10, will a Q10 of two happen approximately for all enzymes?
No, average doesn't mean all enzymes will be approximately doubled in rate of reaction every increase in 10 celsius.
Between 0 and 40 degrees celsius, how are the reactions increased for every 10 celsius increased?
Every 10 celsius the temperature rises, the rate of reaction doubles for most enzymes as more kinetic energy is available for the enzymes and substrates to successfully collide.
What organic compound acids dissociate?
Lactic acid <--> H+ and lactate
Pyruvic acid <--> H+ and pyruvate
What are H+ ions also known as?
Protons
What does a pH buffer do?
Resist changes in pH
Where in the body is there an example of a buffer?
In the blood
What must the pH of the blood be around? What keeps the pH near that value?
pH of 7.4, pH remains narrow towards this due to buffer.
How do hydrogen ions interfere with the tertiary structure of enzymes?
- H+ ions will be attracted to the negatively charged ions/molecules
- H+ ions will interfere with the hydrogen bonds and ionic bonds, tertiary structure of enzyme's active site will change shape
- Substrate molecule will not fit properly into the active site, rate of reaction will be lowered.
What happens if you keep increasing hydrogen ions with enzymes?
- Alter the charges of the active site of enzyme molecules as more hydrogen ions cluster in active site
- Some hydrogen ions will react with certain amino acids
- Bonding between enzyme and substrate will eventually stop completely as enzyme tertiary structure changes completely and substrate can no longer bind to it's active site. Enzyme denatured
What happens if the pH changes slightly on either side of the optimum pH?
Rate of reaction slows down, active site of enzyme slightly disrupted.
Is changing the pH reversible?
Yes. If normal pH is restored and is a slight change, hydrogen bonds can reform and active site shape is restored. Rate of reaction returns to normal
What happens at extreme pH's?
Enzyme's active site become permanently changed. Enzyme is now denatured.
What is a rough estimate of the pH of enzymes working intracellularly?
Around pH 7.
What is a rough estimate of the pH of enzymes working extracellularly?
Different values for pH. There is no average.
What pH does amylase work best at?
pH 6.8
What pH does pepsin work best at?
Between pH 1 and pH 2.
Why does pepsin in the stomach need to work at this pH?
As the stomach secretes hydrochloric acid, with many protons and pepsin has adapts to the low pH.
What type of enzyme is pepsin?
Protease enzyme, hydrolyses large protein molecules into smaller molecules
What pH does trypsin and enterokinase work best at?
pH of 7.8
Why does trypsin and enterokinase work best at this pH?
Salts and bile made from the liver neutralise the pH of the solution from the stomach, protein-digestive enzymes are more adapted to 7.8.
What happens if there is no substrate present in a solution of enzymes?
No reaction can happen. (duh)
What happens if you increase substrate levels with enzymes? (not to the maximum rate just yet)
Rate of reaction increases as;
- More enzyme-substrate complexes can form
- Therefore more product is formed from the product-substrate complex
- Substrate concentration is still the limiting factor as rate of reaction increases to the increase of substrate concentration.
What happens to the rate of reaction as substrate concentration increases indefinitely? (5)
- Maximum rate will be achieved
- Increasing substrate concentration will not increase rate of reaction
- Due to all of the enzyme's active site being occupied with substrate molecules
- Maximum Enzyme-substrate complex's formed
- Substrate concentration is no longer the limiting factor
- Enzyme concentration is now the limiting factor as lack of enzyme molecules prevents rate of reaction from increasing.
