enzymes Flashcards
(101 cards)
1
Q
enzymes are ___________ _________.
A
biological catalysts
2
Q
why are enzymes biological catalysts?
A
enzymes are globular proteins produced in living cells.
exception: a class of enzymes that are RNA in nature
3
Q
why are enzymes biological catalysts?
A
enzymes speed up chemical reactions while remaining unchanged in the process. they do not cause reactions to take place. they merely speed up reactions that would ordinarily proceed, but at a much slower rate in their absence.
4
Q
properties of enzymes (short version)
A
- highly efficient in very small amounts
- highly specific
- reactions are reversible
- easily denatured
- affected by various factors
5
Q
properties of enzymes (long version)
A
- they are highly efficient in very small amounts (average about 1000 reactions per second, 10³ to 10⁶ times faster than uncatalyzed reactions.)
- they are highly specific and thus responsible for its ability to catalyse only certain chemical reactions
- enzyme catalysed reactions are reversible
- enzymes are easily denatured at high temperature/heat and high (basic/alkaline) or low pH (acidic)
- their activity can be affected by various factors such as pH, temperature, substrate concentration, cofactors / coenzymes, enzymes concentration and inhibitors
6
Q
where can we find enzymes?
A
intracellular enzymes:
- in the cytosol or nucleus (nucleoplasm)
- inside membranous organelles e.g. mitochondrial matrix, stroma of chloroplasts, lysosomes
- attached to membranes e.g. plasma membrane, mitochondrial cristae, lamellae of chloroplasts
extracellular enzymes:
- enzymes that are produced in the cell but packaged to be secreted from the cell and work externally. e.g. digestive enzymes
7
Q
why do we need enzymes?
A
without enzymes, the many chemical and metabolic reactions occurring in living organisms will be unable to be carried out at a rate fast enough to sustain life.
8
Q
why can’t humans use heat instead of enzymes to speed up chemical reactions?
A
normal body temperature (5-40 degree celcius) is too low a temperature for chemical reactions to proceed at a fast enough speed to sustain life.
raising the temperature above 40 degree celcius would increase reactions rates but protein would be denatured and lose its unique structure and properties.
enzymes help to promote the reaction and enable it to proceed rapidly at a suitable moderate temperature (i.e. temperature within physiological limits)
9
Q
the chemical reactions that occur within cells are referred to as
A
metabolism / metabolic reactions.
10
Q
the two types of metabolic reactions
A
- anabolic reactions synthesise complex substances, usually using up energy in the process
- catabolic reactions breakdown complex substances into simpler ones, usually releasing energy in the process.
11
Q
what is a metabolic pathway?
A
the formation of a particular substance via a series of intermediate steps, each governed by a specific enzyme.
12
Q
two roles of enzymes in the cell
A
- catalysis: enzymes act as highly specific catalysts that speed up the rate of metabolic reactions
- regulation: enzymes provide a mechanism whereby individual reactions can be controlled; the available quantity of an enzyme determining the rate of the corresponding reaction.
13
Q
list the six major groups of enzymes
A
- oxidoreductases
- transferases
- hydrolases
- lyases
- isomerases
- ligases
14
Q
oxidoreductases - type of reaction
A
oxidation/reduction reactions
15
Q
two types of oxidoreductases
A
oxidases
dehydrogenases
16
Q
transferases - type of reaction
A
transfer a functionally important group from one molecule to another
17
Q
hydrolases - type of reaction
A
split molecules in two by action of water
18
Q
lyases - type of reaction
A
add or remove groups without involving water
19
Q
isomerases - type of reaction
A
convert one isomer of a compound into a different isomer
20
Q
ligases - type of reaction
A
link together two molecules at the expense of ATP
21
Q
what are the 4 categories of amino acids in an enzyme?
A
- catalytic residues
- binding residues
- structural residues
- non-essential residues
22
Q
function of catalytic residues
A
make or break chemical bonds;
the basis of catalytic activity
23
Q
function of binding residues
A
hold substrate in place during catalysis
24
Q
function of structural residues
A
maintain the correct globular shape of the active site so that enzyme can function properly
25
location/function of non-essential residues
near the surface of enzyme; no specific functions thus can be removed or replaced without loss of function
26
what is an active site
a small pocket / groove / cleft on the surface of an enzyme molecule, formed by the precise folding of the enzyme
27
the active site has
a specific 3 dimensional configuration and distribution of charges, which is complementary to that of its substrate
28
the active site typically consists of how many amino acids?
3-12
29
what categories of amino acids are typically in the active site?
catalytic and binding residues
30
function of binding residues in the active site
hold the substrate in the active site by weak interactions between R-groups and substrate e.g. hydrogen bonds and ionic bonds
31
function of catalytic residues in the active site
R-groups help to catalyse the conversion of the substrate to product
32
lock and key hypothesis
- substrate is "key" and enzyme is "lock"
- substrate's shape is complementary to the shape of the active site
- substrate fits exactly into active site of enzyme
- substrate binds to active site of the enzyme, forming enzyme-substrate complex
- once products are formed, they no longer fit into the active site and are released
33
induced fit hypothesis
1. 3D conformation of substrate is still complementary to shape of active site but does not fit exactly.
2. Binding of the substrate to the active site induces a conformational change in the conformation of the enzyme.
3. This enables the substrate to fit more snugly into the active site when they form the enzyme-substrate complex
4. products are formed and released as they no longer fit into the active site
not sure about this one:
This explains why certain enzymes can work on a range of (structurally similar) substrates
34
how does the lowering of activation energy occur in induced fit hypothesis
this induced fit may strain substrate bonds or place chemical groups of the active site so in position to catalyse the reaction. in reactions involving two or more reactants, the active site may hold the substrates in the proper orientation for a reaction to occur
35
activation energy is required to
destabilise existing chemical bonds and to initiate a chemical reaction
36
activation energy is often supplied in the form of
heat energy that the reactant molecules absorb from the surroundings
37
formation of the enzyme-substrate complex increases/decreases the energy of activation of the reaction.
decreases
38
how do enzymes lower activation energy?
enzymes work by lowering the activation energy barrier via re-routing the overall reaction through several steps, each with smaller activation energies. these small steps include the transient formation of an enzyme-substrate complex.
39
molecular mechanisms which contribute to a lowering of activation energy
1. proximity effects
2. strain effect
3. orientation effects
4. microenvironment effects
5. acid-base catalysis
☆ for first three!
40
proximity effects
temporary binding of reactants next to each other on an enzyme increases the chance of a reaction.
uncatalysed reactions depend on random collisions between substrate molecules.
41
strain effects
slight distortion of the reactants as they bind to the enzyme strains the bonds which are to be broken and increases the chance of breakage
42
orientation effects
reactants are held by the enzyme in such a way that bonds are exposed to attack
43
microenvironment effects
hydrophobic amino acids create a water-free zone in which non-polar reactants may react more easily
44
acid-base catalysis
acidic and basic amino acids in the enzyme facilitate the transfer of proton to and from the reactants
45
the effectiveness of acid-base catalysis depends on
the precise position of positive and negative electrical charges on the amino acids of an enzyme.
46
the precise position of positive and negative electrical charges on the amino acids of an enzyme is affected by
the pH of the solution
47
exergonic reaction
chemical products have less free energy than the reactant molecules.
reaction is energetically downhill.
△G is negative
48
endergonic reaction
products store more free energy than reactants.
reaction is energetically uphill.
△G is positive.
49
three steps in the action of enzymes. (simple version)
1. formation of enzyme-substrate complex
2. catalysis/lowering of activation energy
3. product formation and regeneration of enzyme
50
three steps in the action of enzymes (long ver.)
1. Formation of enzyme-substrate complex
· Both the shape and charge of the active site only allow substrates to enter it in specific/proper orientations to put them in a very precise collision.
· A small rearrangement (induced fit) of chemical groups occurs in both the enzyme active site and the substrate molecules.
· Certain binding residues of the active site may temporarily form weak, but extensive bonding with substrates - the enzyme-substrate complex is formed.
2. Catalysis/Lowering of activation energy
· Interactions between the enzyme and substrate molecules occur; straining the chemical bonds within the substrates. Thus, lowering the activation energy.
3. Product formation and regeneration of enzyme
· When the final reaction between the substrates is finished, the product(s) no longer fit properly into the active site and are expelled.
· The temporary changes in shape, charge and bonding pattern within the enzyme revert to their original state, and the enzyme is ready to accept another set of substrates.
51
rates of enzyme-catalysed reactions can be measured by
the amount of substrate changed over time or the amount of products formed over time
52
the actual value of the rate of enzyme-catalysed reaction can be calculated by
measuring the slope of the tangent at the initial part of the curve
53
factors that affect rate of enzyme catalysed reactions
enzyme concentration, substrate concentration, temperature and pH
54
how does increasing enzyme concentration affect rate of enzyme catalysed reactions?
When enzyme concentration is increased, the rate of reaction increases proportionally to the increase in enzyme concentration, provided substrate is in excess. This is because there are more active sites available for substrates to collide with.
If the amount of substrate is limited, a point will be reached when increasing the enzyme concentration no longer has any effect as there are many more empty active sites than there are substrates. Enzyme concentration is no longer the limiting factor for the reaction, and the addition of more enzymes does not alter the rate of reaction. Substrate concentration becomes the limiting factor. Increasing substrate concentration will then increase the rate of reaction.
55
how does increasing substrate concentration affect the rate of enzyme catalysed reactions?
The rate for most enzyme-controlled reactions varies with the availability of substrate.
When enzyme concentration is fixed and substrate concentration is limiting, an increase in the substrate concentration will lead to an increase in the rate of reaction.
As the amount of substrate becomes excessive, the rate becomes more dependent on the amount of enzyme available. The rate of reaction will start to taper off as all the active sites have been filled. This will be the theoretical maximum rate of reaction for that enzyme and is referred to as the Vₘₐₓ.
56
at low substrate concentration, how does rate of reaction change with an increase in substrate concentration?
At low [S], the rate of reaction increases proportionately with increase in [S]. Many available enzyme molecules have active sites, which are unoccupied, and the limited supply of substrate molecules largely determines the reaction rate. Increasing [S] results in an increase in the frequency of effective collisions between substrates and enzymes, and the rate of formation of products, thus increasing the rate of reaction.
57
at high substrate concentration, how does rate of reaction change with an increase in substrate concentration?
At a certain [S], the enzyme active sites become fully occupied/fully saturated because the formation of enzyme-substrate complex is slightly faster than the release of the products. The substrate is said to be in excess, and the rate of reaction becomes constant and shows no increase. The excess substrate molecules are queuing for vacant active sites. An increase in rate can only be achieved by increasing the amount of enzyme available.
58
how does temperature affect the rate of enzyme catalysed reactions?
Temperature affects molecular motion.
At near or below freezing point, enzymes are inactivated (not denatured) as they have very little kinetic energy for effective collisions. At higher temperature, they regain their catalytic influence.
When temperature increases, the kinetic energy of enzymes and substrates increases, thus increasing the frequency of effective collisions between enzyme and substrate causing more enzyme-substrate complexes to form, leading to a greater rate of reaction.
However, the rate of reaction can only increase up to a maximum level called the optimum temperature. Beyond this, further increases in temperature continues to increase the kinetic energy of the enzyme, which eventually leads to a breakage of the bonds (e.g. hydrogen bonds and hydrophobic interactions) that hold the enzyme (protein) in its 3-D conformation. The enzyme is said to be denatured and is no longer effective in catalysing reactions. This is because the 3D configuration of the active site has been changed and the substrate no longer fits into it. Renaturation of enzyme where the enzyme folds back into its original conformation can occur provided that the conformation of the enzyme is not drastically altered.
59
optimum temperature for enzymes in mammals
about 37°C
60
optimum temperature for human enzymes
35-40°C
61
optimum temperatures of enzymes of arctic plants and mammals
about 10°C
62
optimum temperature of some algae in hot springs
(if you're cramming just skip this lmao)
80°C
63
temperature coefficient formula
Q₁₀ = (rate at temperature (t + 10)°C) / (rate at temperature t°C)
64
Typically between 0 to 40°C, the rate of enzyme reaction is ___ for every 10°C increase in temperature. The Q₁₀ is thus ___.
doubled, 2
65
how does pH (& salt concentrations) affect rate of enzyme catalysed reactions?
Enzymes operate optimally over a very narrow pH range. Changes in pH and salt concentrations affect enzymes in 2 ways:
(a) Effect on neutralising charges of amino acids at active site
· An enzymes’ active site comprises of binding and catalytic residues. These are usually charged amino acids that interact with the substrate.
· If pH increases / decreases, the excess OH- / H+ ions combine with the positive / negative charges in the active site, neutralizing them. The uncharged binding and catalytic amino acid residues at active site of enzyme can no longer interact with the substrate and so catalytic activity is lost.
(b) Effect on changing the conformation of enzyme and its active site
· As acidity increases or decreases, the number of H+ ions increase or decrease. This will disrupt the ionic bonds within the enzymes, thus altering the conformation of the enzyme. The 3D configuration of the active site would thus be changed leading to a decrease in the rate of reaction.
· Enzymes become denatured; particularly at extremes of optimum pH values, thus catalytic property is lost. Unlike the effects of heat on enzymes, the effects of pH are normally reversible, at least within limits. Restoring the pH to the optimum level usually restores the rate of reaction.
66
why do enzymes have the ability to be inhibited?
so that cells can regulate their activity.
67
enzyme inhibition may be irreversible if:
Covalent bonds form between inhibitor and enzyme such that inhibitor permanently binds to the active site or alters the shape of active site.
Examples include toxins and poisons
68
enzyme inhibition may be reversible if
- Only weak bonds e.g. hydrogen bonds form between inhibitor and enzyme
- The effect is temporary and causes no permanent damage to the enzyme
- Removal of the inhibitor is possible and restores the activity of the enzyme to normal.
69
what to note in enzyme inhibition
as long as there are available enzymes, the final amount of products made is still the same, as there will still be active sites available for the substrate to enter. The rate of reaction however, is reduced in the presence of the inhibitor.
70
properties of irreversible inhibitors
- Irreversible inhibitors attach to the enzyme at any point, maybe even the active site.
- They usually permanently combine with the sulphydryl groups.
- They will then change the structure of the enzyme and thus the enzyme becomes ineffective. They are thus generally known as poisons. E.g. nerve gas, pesticides, antibiotics... etc.
- Increasing [S] will not reduce the effect of the inhibitor.
71
what are the two types of inhibitors in reversible inhibition?
competitive and non-competitive
72
competitive inhibition
(similarity of inhibitor to substrate, how it inhibits, how it affects rate of reaction)
In competitive inhibition, the binding sites of the inhibitor molecules have a similar shape to the substrate molecule. This means that it is able to fit into the active site of the enzyme. This type of inhibitor thus *competes* with the substrate for the limited enzyme active sites, preventing the substrate from entering the active site, thus reducing reaction rate. It does not alter the shape of the active site.
The inhibition can be overcome by increasing substrate concentration so that as active sites become available, more substrate molecules than inhibitor molecules are around to gain entry to the active sites. i.e. increasing [S] increases rate of reaction. Rate of reaction still reaches maximum / Vₘₐₓ (similar to normal reaction in the absence of inhibitors) but at a higher substrate concentration.
73
example of the use of competitive inhibition in drugs
- Competitive inhibition has been used as the basis for a group of antibiotics called sulphonamides.
- This antibiotic competitively inhibits the pathway involved in the formation of folic acid - an essential amino acid for bacteria.
74
non competitive inhibition
(does the inhibitor bear resemblance to the substrate molecule? where does the inhibitor bind to? what does the inhibitor binding to the enzyme do? how is the rate of reaction affected?)
• The binding sites of the inhibitor bear no resemblance to the substrate molecule. It does not attach itself to the active site of enzyme, but binds to the enzyme at a point other than the active site.
• Binding of the inhibitor to that point alters the conformation of active site such that the substrate no longer fits into the active site.
• When inhibitor concentration increases, the rate of reaction decreases.
• Unlike competitive inhibition, an increase in substrate concentration cannot overcome enzyme inhibition. Rate of reaction reaches a lower maximum level as compared to that without inhibitor.
75
list the measures which regulate enzyme activity in cells
1. compartmentalisation / localisation
2. inactive forms / zymogens
3. allosteric regulation
- 3A. end-product inhibition / negative feedback inhibition (allosteric inhibition)
- 3B. allosteric activation
4. regulation of enzymes by phosphorylation
5. genetic control: induction and repression of enzyme synthesis
76
compartmentalisation / localisation
(what it is + its function)
many enzymes are localised in specific organelles within a cell. such compartmentalisation serves to:
- isolate the reaction substrate or product from the competing reactions
- provide a favourable environment for the reaction. e.g. low pH in lysosomes
- organise the thousands of enzymes present in the cell into purposeful pathways. E.g. respiratory enzymes in mitochondrion
77
inactive forms / zymogens
(what exactly it is when regulating enzyme activity + example)
Cells may synthesize enzymes in their inactive form. Pepsin is a powerful protein-digesting enzyme capable of disrupting a cell’s internal structure. The stomach cells produce pepsinogen, the inactive form of pepsin which becomes active only when exposed to strong acid conditions.
78
what to note when using the term "allosteric regulation"
only use it if question stated "allosteric site", "allosteric enzyme", etc.
79
most enzymes having allosteric sites are
proteins comprising two or more polypeptide chains/subunits. Each subunit has its own active site and allosteric sites which are usually located where subunits are joined.
80
allosteric enzyme can exist in two conformational states:
- one catalytically active
- the other inactive
81
how does allosteric inhibition affect the rate of reaction? (like as in how the allosteric inhibition works to affect the rate of reaction + what is the affect)
binding of an allosteric inhibitor to the allosteric site causes a change in the conformation of active site of the enzyme. The enzyme in its inactive form and the substrate can no longer bind with the enzyme. This reduces the rate of reaction.
82
how does allosteric activation affect the rate of reaction?
binding of an activator to the allosteric site of an enzyme stabilises the conformation of the active site of enzyme. this increases the rate of reaction.
83
function of end-product inhibition / negative feedback inhibition
provides a dynamic and flexible way to self-regulate enzyme pathways.
84
describe end-product inhibition / negative feedback inhibition
• The enzyme catalysing the first step in a biochemical pathway often has an inhibitor-binding site (allosteric site) to which the end-product produced at the last step of the series of chemical reactions binds. When the final product of a sequence of reactions starts to accumulate, the enzyme pathway leading to its manufacture can be shut down when it is no longer needed.
• The final product acts as an allosteric inhibitor for the enzyme at the beginning of the pathway i.e. the activity of the enzyme is limited because the product molecule binds to an allosteric site of enzyme.
• In this type of inhibition, the final product itself inhibits the enzyme producing it. This serves as a kind of self-regulation, preventing over-production of the product.
• This is a reversible process. As the product is used up, the inhibition is lifted and the production of end-product is switched back on.
85
is end-product inhibition / negative feedback inhibition reversible? why?
Yes, this is a reversible process. As the product is used up, the inhibition is lifted and the production of end-product is switched back on.
86
describe regulation of enzymes by phosphorylation
• A family of enzymes, called protein kinases, catalyses phosphorylation reactions. These kinases use adenosine triphosphate (ATP) as a phosphate donor to add phosphate groups to enzymes.
• Phosphate groups are cleaved (removed) from phosphorylated enzymes by the action of phosphoprotein phosphatases.
• Depending on the enzyme, the phosphorylated form may be more or less active than the unphosphorylated enzyme.
87
example of regulation of enzymes by phosphorylation
For instance, phosphorylation of glycogen phosphorylase (glycogen degradation) increases activity, whereas the addition of phosphate to glycogen synthase (an enzyme that synthesizes glycogen) decreases activity.
88
describe genetic control: induction and repression of enzyme synthesis
• Cells can regulate the enzymatic activity of the cell by altering the rate of enzyme synthesis.
• Alterations in enzyme levels due to induction or repression of protein synthesis are slow (hours to days) compared to allosteric changes in activity which occur in seconds to minutes.
89
what are enzyme cofactors?
they are non-protein substances required for the efficient functioning of an enzyme.
90
characteristics of cofactors
• Enzymes ‘helpers’ (promote enzyme activity) but have no catalytic properties of their own
• Non-protein; thus stable at relatively high temperature
• Smaller than the enzymes with which they are associated.
• May undergo a temporary change during reaction, but they are normally restored afterwards therefore reusable.
• Required in small amounts
91
enzyme (apoenzyme) + cofactor =
holoenzyme
92
apoenzyme refers to
the protein portion of the holoenzyme
93
list the three types of cofactors
- inorganic ions
- coenzymes
- prosthetic groups
94
how do inorganic ions function (as a type of cofactor)
They are believed to mould the enzyme into the required shape for the enzyme/substrate complex to be formed.
95
inorganic ions (type of cofactor) is also called
enzyme activators
96
examples of inorganic ions (type of cofactor)
Cl- and salivary amylase; Mg2+ and ATPase
97
coenzymes are derived from
water soluble vitamins, e.g. vitamin B₆
98
what are coenzymes, and how do they usually function?
they are organic molecules which bind loosely and briefly to enzymes and usually act as a hydrogen acceptor or electron carrier
99
example of coenzymes
Nicotinamide Adenine Dinucleotide (NAD)
Important H carrier in biochemical systems, involved in respiration.
NAD (oxidised form) can exist in reduced form (NADH)
100
what are prosthetic groups?
these are non-protein groups that are covalently bonded (tightly bounded) to enzymes. they do not dissociate from the enzyme and assist in the catalytic function of the enzyme.
101
example of prosthetic group
Haem
An iron containing prosthetic group which can be an electron or oxygen carrier. Other than its role in oxygen transport in haemoglobin, haem is also the prosthetic group of cytochromes which play an important role in respiration.
