enzymes Flashcards
Enzymes:
globular proteins with tertiary structure, acting as a biological catalyst. They speed up chemical reactions by lowering the activation energy (Ea). Tertiary structure (3D) provides a specific shape of active site. Needed in small quantities and can be reused.
Activation Energy:
minimum energy required to start the reaction, make new bonds in product, break bonds in substrate
▪ Can be lowered by using heating or using enzymes
what makes an enzyme specific?
- specific sequence of amino acids in primary structure
- determine the arrangement and the type of R groups
- which determines the type of bonds and interaction between R groups
- Thus in turn, it determine the overall folding and coiling of the polypeptide,
giving a precise 3D shape ( tertiary structure ) which brings amino acids close together
So enzymes have specific shape of active site
Structure of Enzyme
▪ Globular protein, 3D shape, tertiary structure
▪ Has active and catalytic site
▪ Made up of many amino acid but only few become part of the active site and others become part of catalytic site
Protein property
Globular protein
Tertiary structure - provide specific shape
Provides shape of active site where reaction occurs
Influence by temperature and pH at extreme of temperature and pH, they denature
Catalytic property
Remains unchanged at the end of reaction/ can be reused
Increases the rate of reaction
Reduces activation energy
specific
Site of action of enzymes
All enzymes are proteins that are produced via the process of protein synthesis inside cells , some enzymes remain inside cells, whilst others are secreted to work outside of cells. Enzymes can therefore be intracellular or extracellular, referring to whether they are active inside and outside the cell respectively
▪ Intracellular enzymes are produced and function inside the cell
▪ Extracellular enzymes are secreted by cells and catalyze reactions outside cells ( example: digestive enzymes in the gut). Some organisms secrete enzymes outside their bodies. Fungi, for example, often do this in order to digest the food on which they are growing.
Types of enzymes based on metabolic reaction
▪ Catabolic - breaking of large molecules into small
Anabolic- building up of large molecules from small one
2 main approaches to investigate if a reaction took place
▪ Disappearance of reactants
Appearance of products
How do enzymes catalyze specific reactions? (4m)
Active site has specific shape ( in reference to the active site ) which is complementary to the substrate combine to form enzyme-substrate complex, substrate alters shape of the active site, induced fit. Reference to temporary bonds (Hydrogen, ionic and disulphide)
Active site
the site on the surface of enzymes with a particular shape into which only one type of substrate can fit. It is represented by 3-12 amino acids, it consists of R groups which are free to make bonds with substrate
Allosteric site
amino acid which is not a part of active site
process of ESC
Process
1) Substrate enters the active site: enzyme changes shape such that its
active site enfolds the substrates( induced fit)
2) Substrates held in weak interactions, such as hydrogen bonds and ionic
bonds
3) Active site can lower activation energy and speed up the reaction
4) Substrate is converted to products
5) Products are released
Active site is now available for new substrate
How are enzymes soluble in cytoplasm?
Globular proteins, enzyme molecules are coiled into a 3D shape, hydrophilic R groups ( side chains) on the outside of the molecule to make them soluble
Substrate
molecules that bind to the enzyme
Lock and key hypothesis:
ypothesis for enzyme action, the substrate is a complementary shape to the active site of the enzyme and fits exactly into the site , the enzyme shows specificity to the substrate
▪ Each enzyme will act on only one type of substrate molecule . This is because the shape of the active site will only allow one shape of the molecule to fit. The enzyme is said to be specific for this substrate.
The substrate is held together in place by
temporary bonds which form between the substrate and some of the R groups of the enzymes amino acids, the combined structure is called the ES complex.
Induced fit hypothesis (more flexible)
a hypothesis for enzyme action, the substrate is a complementary shape to the active site of the enzyme, but not an exact fit - the enzymes or sometimes the substrate can change the shape slightly to ensure a perfect fit, but is still described showing specificity
Mostly same as lock and key hypothesis buy it adds the idea that sometimes the substrate molecules enters the enzyme in order to ensure a perfect fit. This makes the catalysis even more efficient
How an enzyme catalyzes the breakdown of a substrate molecule into 2 product molecules
an enzyme has a cleft in its surface, called the active site. the substrate molecule has a complementary shape
the interaction of the substrate with the active site breaks the substrate apart . an enzyme-product complex is briefly formed before the two product molecules leave the active site, leaving the enzyme molecule unchanged and ready to bind with another substrate molecule
random movement of enzyme and substrate brings the substrate into the active site. an enzyme- substrate complex is temporarily formed. the R groups of the amino acids in the active site interact with the substrate
the overall rate of reaction for both locke and key and induced fit is
very high
In this example: the enzyme is catalyzing the joining together of 2 molecules
factors affecting enzyme action
1)ph
2) enzyme concentration
3) substrate concentration
4) temperature
how does ph affect enzyme con
it measures the concentration of H+ in a solution. Most Enzymes work in a narrow range of ph. At Optimum Ph Enzymes show its maximum activity above or below the optimum pH enzyme activity decreases. Change in Ph breaks ionic and hydrogen bond between the R groups, it changes the 3D shape of enzymes. Active site of enzyme is deformed. Substrate can no longer fit into the active site. Reaction of reaction slows down and stops
enzyme graph ph
enzyme con graph
As the enzyme con. Increases, there’s an increase in con. After x has
no effect on rate and it remains constant at y, rate becomes plateau
describe region A
As the concentration increases , rate of reaction increases gradually at an enzyme con. Of x, the rate of the reaction was its maximum y
explain region A
With more enzyme molecules there are more molecules , there are more active site available for the substrate to fit. Many enzyme substrate complex formed/ more product. Limiting factor- enzyme, con. substrate are free
describe region B
As the enzyme is above the value of km constant at y
explain region B
All the substrate have formed enzyme complex. Enzyme con. Is no longer the limiting factor but the substrate con. Is the limiting factor, active site are free
As substrate con. Increases, rate of reaction increases,
After a point there’s an increase in substrate
concentration has no effect on rate and the curve
Reaches a plateau
describe region A
As the con. Of the substrate increases, the rate of reaction, gradually increases, after point X, it has no effect on the rate of reaction at y which is at its max
explain region A
With more sub molecules of them can fit into the free action sites, more enzyme substrate complex is formed, with more product being formed , rate increases, sub. Con is limiting factor
describe region B
As the con. Of substrate increases above X, rate of reaction remains constant at y
explain Region B
All the enzymes have formed enzyme substrate complex, substrate con. Is no longer the limiting factor but enzyme con. Is, there are no free active site substrate to bind
increase in the temperature increases, the rate of reaction, it reaches
Its maximum optimum temperature, above the optimum temperature,
Above the optimum temperature, the rate of reaction decreases and
Become 0
describe region A
As the temperature increases, rate of reaction a, it reaches its max optimum temperature
explain region A
Increasing the temperature provides more heat energy, it allows enzyme and substrate to attain more KE. Which helps them to move fast and result in more effective collisions, more Enzyme substrate complex formed, more product formed, rate of reaction increases
Describe region B
As the temperature increases above the optimum temperature, rate of reaction decreases and become 0
explain region B
As the temperature increases above the optimum. More neat energy is given, KE increases, bond holding the tertiary structure of protein ( Hydrogen bonds and ionic bonds), it changes the 3D shape of enzymes and active site changes shape, substrate no longer fit into the active site. No enzyme substrate complex, no product and enzyme is said to be denatured
Michaels Km constant
It is the substrate concentration at which an enzyme works at held its maximum rate( half vMax). It is the measure of affinity of the enzyme for its substrate, at this point held the at the active site of the enzyme is occupied by the substrate
Lower the Km value
higher the affinity of the enzyme for its substrate , lower concentration of substrate is required to reach half Vmax
Higher the Km value
lower the affinity of the enzyme
vMax gives information about the
maximum rate of reaction and Km measures the affinity
significance of enzymes
▪ An enzymes performance for its substrates concentration be compared quantitatively
▪ The performance of same enzyme in different organisms can be compared
Inhibitor
substances that reduces the rate of the reaction
Competitive inhibitor has a similar shape to the substrate
and Complementary to the active site
competitive inhibitor
The inhibitor binds with the active site to form an enzyme Inhibitory complex, which stops the substrate from entering The active site, few enzyme-substrate complexes are formed, Initial rate of reaction is low. Increasing the concentration of substrate increases the rate of the reaction
Non- completive inhibitor
▪ Inhibitors binds with allosteric site, changes the shape
Of the active site, substrate is no longer able to bind with active site,
low vMax, high Km, permanent binding
End product inhibition
▪ Type of noncompetitive inhibition, when the concentration of products is high It acts as an inhibitor for any enzyme in the sequence, it binds to the allosteric site, when The concentration of product decreases, the enzyme is switched on
Turns over number:
number of substrate molecules transformed into products per minute
Immobilization of enzyme
Process of retaining/ trapping enzyme to an inert medium
types of immobilization enzymes
1) absorption enzymes
2) covalent bonding
3) encapsulation
absorption enzymes
surface phenomena where enzymes are attached to the surface of materials like Glass
covalent bonding
binding enzyme to cellulose with help of covalent bonding
Encapsulation:
encapsuled in a partially permeable membrane which allows substrate to enter and product to leave
advantages of immobilization enzymes
1) Product not contaminated, can be collected easily
2) Enzymes can be retained/ reused
3) Longer shelf life
4) Enzymes more stable at higher temperature, extreme pH
Able to obtain more product per time
disadvantages of immobilization enzymes
1) Expensive
2) Not efficient
Enzyme at the Centre of bead is less likely to bind with substrates
When a substrate with complimentary shape fits into the active site
A temporary hydrogen/ionic/disulphide bond is formed between the R group of the enzymes amino acid and the substrate
esc conc over time
starts off high and then falls to zero
protease will remove/clean
deposits of enzymes
when it comes to inhibitors
look at vmax! (high conc of inhibitor = even
lower ror
optimum temp go for a
range
ctivation energy required is greater =
ror is reduced or slower!
inhibition is involbed in the regulation of
rate of enzyme-catalysed
reactions
at the peak of ror,
ionic bonds begin to break
product cpnc does not decrease,
it reaches a constant
add substrate to compeitive inhibitor —>
ror increases
disulphide bonds are the last to
break
quarternary also controls the shape of
the active site
all 4 protein structures can determine the specificity
of an enzyne and all 4 control the shape of the active site but only primary, secondary and tertiary must be involved in the
formation of an active site
if an enzyme gets denatured at 50,
eaction stops before then only tertiary structure is always involved when comp and non comp inhibitor bind to enzymes
denaturation does not involve
the primary structure
comp inh dont have exactly the same shape as substrate, i
it is just
SIMILAR
Enzyme w the lower vmax uses more of the substrate
cuz it has higher
affinity and lower km (draw out a graph)
What is meant by specificity of enzyme
Enzyme acts on only one substrate
Shape of active site complementary to substrate
How does enzyme lower Ea: activation energy
Provides alternative energy pathway
Brings reactants close together to from ESC
Puts strain on reactant
So bonds break easily
Transfer of charges between groups
Changes orientation such that reaction is feasible
How to describe and explain enzyme results (D = description, E = explain)
E - not all active site occupied
E - so substrate concentration is limiting
E - so fewer ESC formed
D - at high concentration less steep increase in rate of reaction
D - it levels out/plateaus at (XYZ) concentration
E - enzyme concentration is limiting
E - since all active sites are occupied/saturated
E - Vmax reached
Advantages of enzymes with high optimum:
The production/process requires high temperature
High temperature so more collisions and more ESC
Higher ROR increases
More stable and can be used again and again
Less prone to denaturation
Works well over a range of temperatures and pH so works at higher rate
Outline experiment that should be carried out to find out if inhibitor is competitive or non-competitive:
Carry out experiment with and without an inhibitor
At many different concentration of substrates
Keeping other variables constant (eg pH and temp)
Draw a graph of rate of reaction against substrate concentration
If inhibitor is competitive than Vmax is same as without inhibitor
Describe mode of action of enzyme with induced fit
Active site changes shape OR moulds around substrate OR better fit
Active site becomes fully complementary to substrate
Formation of ESC
Lowers activation energy
By providing alternate energy pathway/puts strain on bonds
Breaking of bonds
Active site returns to initial stage AND can be reused
After XYZ substance/device is used why does enzyme B have a lower Km and why does enzyme C have a higher Vmax
have a higher Vmax
B: increases affinity of enzyme for substrate
B: makes shape of active site more complementary
B: Makes position of active site more accessible
C: increase rate of ESC formed
C: increases rate of catalysis after binding
C: may lower activation energy required than normal
Advantages of end-product inhibition when enzyme in intracellular:
Maintains balance
Efficient metabolism
Avoids osmotic problem that could be caused by build up of product
Disadvantages of end-product inhibition when the enzyme is used extracellular and commercially
Loss of product
Slow rate of reaction
Product required continuously
Why are immobilised enzymes more active over range of pH
Alginate covering is protective so the H+/OH- ions do not penetrate alginate beads so the shape of the active site is not disrupted.
Explain what is meant by higher Km
Enzyme has lower affinity for substrate
Needs a higher concentration of substrate to reach Vmax
Less likely to be saturated by substrate
Suggest one other advantage of using enzymes obtained from microorganisms, rather than enzymes extracted from barley seeds, in the production of sugar syrups.
easier to extract
idea that microorganisms can be cultured in large quantities and produce large amounts of enzyme
higher rate of reaction
active over a greater temperature range
Advantages of immobilised enzymes compared to enzymes in free solutions:
(may be able to) obtain more product (per unit time) ;
can use higher temperatures (to obtain more product) / still active at higher temperatures ; A thermostable
does not denature (as easily as free) if temperatures increase / AW ;
enzyme can be easily recovered ;
downstream processing is easier ;
product, not / less, contaminated ; A less purification needed
longer shelf-life of enzyme ;
reduces product inhibition ;
enzyme is, more stable / less likely to denature or described ; A thermostable / can work at high temperatures A in context of change in pH I ‘can withstand changes in temperature’
Explain how the addition of the competitive inhibitor results in the same value for Vmax but a higher value for Km.
Competitive inhibitor occupies the enzyme’s active site / competes with substrate
Reduces frequency of successful collisions / fewer ESC formed
Reduces reaction rate at low substrate concentration
idea that curve with inhibitor is to the right of the curve without inhibitor ;
At high substrate concentration, the effect of inhibitor is reduced
Therefore the Vmax is the same as it is determined by the concentration of enzyme
A explanation in terms of active sites, saturated / fully occupied
idea of intercept to curve gives a higher value for Km ;
