Enzymes Flashcards
What is an enzyme?
A protein molecule that acts as a biological catalyst by lowering activation energy
what type of protein is an enzyme and how does it help?
globular protein so hydrophillic amino acids on outside and hydrophobic on inside - makes it soluble in water
anabolic reaction
builds up molecuels
catabolic reactions
break down molecules
metabolism
combination of anabolic and catabolic reactions - all different reactions happening in a cell
metabolic pathway
sequence of enzyme controlled reactions
extracellular
enzymes that catalyse reactions outside of cells eg. breaking down nutrient molecules for digestion
intracellular
enzymes that catalyse reactions inside cells eg. synthesising polymers from monomers like making polysaccharides
example of intracellular enzymes and what it breaks down
- catalase - hydrogen peroxide to water and oxygen
examples of extracellular enzymes and what they break down
- Amylase (produced by salivary glands and pancreas) - starch into maltose
- Trypsin (produced by pancreas) - protein into peptides
structure of an enzyme’s active site
amino acids interact with each other to maintain a specific tertiary structure - means it has a specific active site to catalyse specific reactions
induced-fit hypothesis
- the tertiary structure of the active site is flexible and changes shape slightly as the substrate enters
- the bonds formed between substrate and enzyme help catalyse the reaction, lowering activation energy
- when the product leaves the enzyme the active site returns to its inactive state
- other substrate molecules cannot form the correct bonds with the active site so the tertiary structure doesn’t change shape
lock and key hypothesis
- a specific substrate binds to the active site of a specific enzyme by interacting with R groups of the active site forming enzyme-substrate complex
- substrate reacts and products formed in an enzyme-product complex
- product released
what can the quaternary structure of an enzyme mean?
- they can have more than one active site
- eg. catalase has 4 identical polypeptide chains and therefore 4 active sites
activation energy
amount of energy that must be applied for the reaction to proceed
how do enzymes affect activation energy?
they lower it
Digestion of starch
- starch polymers broken down into maltose by amylase (produced by salivary glands and pancreas)
- Maltose broken down into glucose by maltase (produced by small intestine)
- glucose is then small enough to be absorbed by cells lining the digestive system
Digestion of proteins and where is enzyme produced?
- Trypsin is a protease that catalyses digestion of proteins into smaller peptides
- then broken down further into amino acids by other proteases and absorbed by cells lining the digestive system
- Trypsin - produced in pancreas and released into small intestine
Effect of low temp on enzyme activity
low rate of reaction
- low kinetic energy so move slowly
- few successful collisions between enzyme and substrate
effect of optimum temp on enzyme activity
high rate of reaction
- lots of kinetic energy so move quickly
- lots of successful collisions between enzyme and substrate
Effect of very high temp on enzyme activity
low rate of reaction
- heat causes loads of kinetic energy and enzymes vibrate very fast - breaks the hydrogen bonds maintaining tertiary structure of enzyme
- active site changes and substrate doesn’t fit
- enzyme denatures
- no successful collisions
what is temperature coefficient (Q10)?
- tells you how much a rate of reaction increases with a 10 degree rise in temp
- measure rate of reaction at certain temp (eg. 20 degrees)
- measure rate at 10 degrees higher (30 degrees)
- temperature coefficient = higher (30)/lower (20)
what does a graph measuring effect of temp on rate of reaction look like?
starts flatter, curves up u-shaped, then rapidly falls down
effect of NOT optimum pH ( too high/low conc. hydrogen ions) on enzyme activity
low rate of reaction
- hydrogen ions can bond with R groups on amino acids of protein (including on active site)
- Hydrogen ions bonding to active site can prevent R groups bonding to substrate, by breaking tertiary structure
- substrate can’t bond with active site
- no successful collisions
graph showing effect of pH on enzyme activity
bell shaped, with highest point changing depending on optimum pH
What effect does low substrate conc have on enzyme activity?
low rate
- few substrate molecules limits chances of successful collisions
what effect does high substrate conc have on enzyme activity?
high rate
- more substrate molecules increases chances of successful collisions
what does graph showing effect of substrate conc look like?
- substrate conc is directly proportional to rate of reaction at first so straight diagonal line through 0
- then reaches Vmax where every enzyme is occupied so graph goes horizontal
What is Vmax?
the maximum rate of reaction
- occurs when all the active sites are occupied by substrates, no more enzyme-substrate complexes can be made until products are released
- enzymes are saturated
- occurs when increasing substrate conc.
effect of low enzyme conc on rate of reaction
low rate
- low chance of successful collisions between enzyme and substrate
what does graph showing effect of enzyme conc look like?
- enzyme conc is directly proportional to rate of reaction at first so straight diagonal line through 0
- then line goes horizontal if substrate conc is not increased becuase there aren’t enough substrates to bind with enzymes
How do you make different concentrations of catalase?
Serial dilution
- add 1ml of stock solution to 9ml distilled water
- add 1ml of that solution to 9ml of distilled water
- repeat a number of times to get serial dilution
method of investigating effect of substrate conc on enzyme activity
- Put 100cm^3 hydrogen peroxide and one potato cylinder into conical flask
- set up conical flask with bung going into upside-down test tube in trough full of water
- record volume of gas given off every 30s for 3 min
- Repeat for other concentrations using 100cm^3 each time
- calculate mean, standard deviation and rate of gas production
How do competitive inhibitors work?
- a molecule a similar shape as a substrate fits into the active site
- This blocks the substrate from entering the active site, preventing the enzyme catalysing the reaction
- slows the rate of reaction
- eg. malonate is inhibitor competing with succinate and can inhibit respiration
examples of competitive inhibitors
- statins - inhibit synthesis of cholesterol for reducing cholesterol conc
- aspirin - irreversibly inhibits enzymes helping make chemicals for producing pain
how do you reduce the effect of a competitive inhibitor?
increase substrate conc. - makes it much -more likely enzyme will collide with substrate rather than inhibitor
how does a graph increasing substrate conc. change when adding a fixed conc. of a competitive inhibitor?
- will go from straight diagonal line, then reaching Vmax and going horizontal to one straight diagonal line reaching the same end point but not going horizontal
what effect do competitive inhibitors have on rate of reaction?
- they slow it down
- much more effective initially but as substrate conc. increases they become less effective
How do non-competitive inhibitors work?
- inhibitor binds to enzyme at the allosteric site
- causes tertiary structure of enzyme to change - active site changes
- active site no longer has complementary shape for substrate to bind to
- cannot form enzyme-substrate complexes lowering rate of reaction
- eg. cyanide
why does increasing the substrate conc not reduce the effect of non-competitive inhibitors?
it causes the shape of the active site to change so even when more substrate is being added there is no active sites for it to bind to
how does a graph increasing substrate conc. change when adding a fixed conc. of a NON-competitive inhibitor?
It is parallel to it but lower down (r-shaped) as increasing substrate conc doesn’t effect it
what is end product inhibition?
the final product in a metabolic pathway inhibits an early stage enzyme in the pathway
- used to reduce rate of metabolic pathway if less product is needed
- example of negative feedback - keeps level of molecules in a set range
- example of non-competitive inhibition
examples of end product inhibition
- eg. in protein synthesis if level of amino acids get too high they will inhibit the first enzyme in the pathway that makes them
- eg. in respiration if ATP levels get too high they inhibit the first enzyme in the pathway that makes them
reversible inhibitors
- bind temporarily to enzyme
- form weak H/ionic bonds with enzyme
- effects reversed by change in environment for enzyme
non-reversible inhibitors
- binds permanently with enzyme
- strong covalent bonds with enzyme
- cell must produce more of the enzyme by activating gene so the enzymes are transcribed and translated
Difference between cofactor and coenzyme
cofactor - a non-protein inorganic molecule that is required by an enzyme to function eg. amylase requires chloride ion
coenzyme - an organic molecule required by an enzyme to function eg. coenzyme A is derived from vitamin B5
- help transfer atoms or groups from one reaction to another or may form a part of the active site
where do we obtain cofactors?
- via the diet through minerals eg chloride, iron, calcium, zinc ions
example of a cofactor
chloride ion is a cofactor needed for amylase to digest starch
When can a co-factor be a prosthetic group?
- prosthetic groups are cofactors - must be tightly bound and forms
a permanent feature of the enzyme
eg. zinc ion is prosthetic group for carbonic anhydrase to metabolise CO2
why is precursor activation needed?
- many enzymes produced in the inactive form - inactive precursor enzymes
- enzyme may cause damage to cells producing them or tissues where they’re released
- precursor enzymes need to undergo a change in shape (tertiary structure) of the active site to be activated
What is the process of precursor activation?
- enzyme is called an apoenzyme
- co-factor added and enzyme is activated - holoenzyme
- change in tertiary structure can also be brought about by the action of another enzyme
what are enzymes that can be activated by a change in environement rather than a cofactor called?
zymogens or proenzymes
how do you measure rate of reaction from a graph showing amount of product in a certain time?
- draw a tangent on the r-shaped curve touching the time we’re interested in
- then calculate the gradient by making it into a triangle and measure vertical side (y) and horizontal side (x)
- y/x
how would you describe the r-shaped graph showing the amount of product made in a certain time with a fixed substrate conc?
(initial, middle, end)
- rapid initial rate - lots of substrate molecules so high frequency of successful collisions
- slower rate - some of the substrate has been turned into product so chance of substrate colliding with active site decreases
- reaction stops - all substrate molecules converted into product so no chance of successful collision
