WK 8 and 9: Enzymes Flashcards
(20 cards)
what is the main role of enzymes and why are they important
catalyse and promote sequences of chemical reactions.
- proteins that catalyse biochemical reactions by lowering their activation energy
- almost every biochemical reaction is catalysed by an enzyme
- under biologically relevant conditions, uncatalyzed reactions tend to be slow (e.g. digesting food would take a long time if not catalysed)
- most enzymes are proteins (with exception of RNA molecules: ribozymes)
- Function of enzymes is to lower the activation energy (Ea) to accelerate the reaction
- enzymes DO NOT affect equilibrium of the reaction. equilibrium is determined by thermodynamics (change in gibbs)
- Enzymes catalyse biochemical reactions by binding to substates. Substrates bind to the region call the active site.
What is a pathway and what is a catabolic and anabolic pathway. what is metabolism
Pathways: consecutive reactions catalysed by enzymes. The products of one reaction become the reactants of the next one
Catabolic pathways: pathways that degrade organic nutrients into simple end products to extract chemical energy and convert it into a form useful to the cell (e.g. ATP, NADH, NADPH, FADH2)
Anabolic pathways: pathways that start with small precursor
molecules and convert them into more complex molecules (e.g. proteins, nucleic acids). These require the use of energy.
What is metabolism and its function
Metabolism: the overall network of enzyme catalysed pathways (anabolic & catabolic)
Overall, the three main purposes of metabolism are:
1) conversion of food to energy to run cellular processes;
2) conversion of food/fuel into building blocks for proteins, lipids, nucleic acids, and carbohydrates;
3) elimination of waste products.
These enzyme-catalysed reactions allow organisms to grow and reproduce, maintain their structures, and respond to their environments.
what are some applications of enzymes
Food: amylase, lactase, protease–> bread, cheese, lactose free milk
Medicine: streptokinase, Asparaginase–> Clot lysis, leukemia treatment
Diagnostics: Glucose oxidase, HRP, Urease–> Blood sugar test, ELISA, H. pylori detection
Therapeutics: CRISPR-Cas9, Prodrug-activating
enzymes, Peptidases and kinases–> Gene editing, site-specific cancer treatment,
Faulty enzymes are targeted in cancer therapy to
disrupt signaling pathways
Cancer therapies that target enzymes
Many anti-cancer drugs target faulty enzymes like protein kinases involved in cancer cell survival and proliferation.
These therapies are referred to as targeted therapies, and have improved efficacy and reduced side effects compared
to traditional chemotherapy.
Imatinib is used to treat patients with Chronic Myeloid Leukemia
- Nomenclature - how name often relates to their function.
- Classified according to type of reaction they catalyse e.g. LDH is an oxidoreductase (Lactate Dehydrogenase)
- Names end in ‘-ase’ and often describe the substate or reaction type e.g. amylase- breaks down starch, lactase breaks down lactate and streptokinase breaks down blood clots
- Structure & function of enzymes.
Function: stabilising the transition state, lowering activation energy, accelerating reaction
- Have an active site: provides environment that helps accelerate reaction
- Binds and recognises specific substrates
E + S ⇌ ES ⇌ EP ⇌ E + P
3 main features that make a protein an enzyme:
1. Catalytic power: accelerate the reaction
2. Specificity: regonises a particular substrate
3. Regulation: control of the enzyme activity by inhibiters or activations
What is catalytic power
accelerates the rate of a biochemical reaction
What is specificity
recognises a particular substrate.
Enzymes act as a template for the reaction to occur, they bind to specific substrate(s), stabilise the transition state, lower the activation energy and accelerate the reaction
Models of enzyme specificity:
1) Lock & key (The active site of an enzyme precisely fits a specific substrate- assumes a rigid conformation)
2) Induced fit model: active site of an enzyme changes the conformation to improve the fit and the binding
* Changes in the active site occur when the substrate comes in the vicinity of the active site
* At the transition state there is a rearrangement of the bonds
* The enzyme wraps around the substrate to facilitate the chemistry of
the reaction
What is regulation
can be regulated by inhibitors or activators.
Inhibitors: inhibit the activity (i.e. interfere with the kinetics of the reaction catalysed by the enzyme)
Activators: increase the activity of the enzyme
TYPES OF INHIBITORS
Reversible inhibitors: interact with enzymes via non-covalent bonds
1)Competitive inhibitor = competes with the substrate for the active site of an enzyme. If the inhibitor occupies the active site, the substrate cannot bind
2)Noncompetitive inhibitor = binds to a site, distinct from the active site, before or after the substrate is bound
to the active site. Binds to a different site, it can bind to the E or to the ES complex.
3) Uncompetitive inhibitor = binds to a site, distinct from the active site, but binds after the substrate is bound to
the active site. binds only to the ES complex.
IRREVERSIBLE: interact with enzymes via covalent bonds to form a stable intermediate. bind covalently with, or destroy, a functional group that is essential for the enzyme’s activity, or they form a highly stable noncovalent association.
regulation: inhibitors
Allosteric inhibitor: induces a conformational change that reduces the enzyme’s affinity for its substrate. Noncompetitive and uncompetitive inhibitors are examples of allosteric inhibition. Vmax is reduced
Allosteric activators: bind to locations on an enzyme away from the active site, inducing a conformational change that increases the affinity of the enzyme’s active site(s) for its substrate(s).
Vmax is increased
ACTIVE SITE, COFACTORS AND HOW ENZYMES WORK
The active site, cofactors, and how enzymes work:
✓ Active site: It provides a specific environment that helps accelerate the reaction; It binds and recognises a specific substrate(s);
✓ Cofactors: a non-protein chemical component needed for the biological activity of an enzyme;
✓ How enzymes work: Enzymes act as a template for the reaction to occur, they bind to specific
substrate(s), stabilise the transition state, lower the activation energy and accelerate the reaction.
WHAT ARE COFACTORS
under specificity
Cofactors:
* Cofactor = non-protein chemical component needed for enzyme activity.
o Inorganic: Fe²⁺, Mg²⁺.
o Coenzyme (organic, loosely bound): NADH, vitamins.
o Prosthetic group (tightly bound): FAD in flavoproteins.
* Apoenzyme = enzyme without its cofactor (inactive).
* Holoenzyme = enzyme with cofactor attached (active).
WK 9 Kinetic Terminology: Michaelis-Menten mode
Michaelis-Menten model:
The rate of an enzymatic reaction depends on the formation of enzyme-substrate complex (ES), which depends on the concentration of substrate, until all the enzyme binding sites are occupied with substrate.
WK 9 Kinetic terminology V₀ (Initial rate):
V₀ (Initial rate):
The reaction is faster at the beginning when there is more substrate and is determined by variation of product over time at the beginning of the reaction.
WK 9 Kinetic terminology Vmax (Maximum rate):
Vmax (Maximum rate):
“Maximum rate achieved in the reaction (when the enzyme is saturated with substrate)”
WK 9 Kinetic terminology Kₘ (Michaelis constant):
Kₘ (Michaelis constant):
“Concentration of substrate, at which the reaction rate is half of Vmax”
- Use of Linear plots to determine kinetic parameters
Lineweaver-Burk plot:
“To linearise the Michaelis-Menten plot, an approach that takes the reciprocal of both sides of the Michaelis-Menten equation”
These plots are used to:
“Determine kinetic parameters and type of enzyme inhibitors”
This includes identifying Vmax, Km, and distinguishing between competitive, noncompetitive, and uncompetitive inhibitors based on changes in slope and y-intercept.
- Types of Enzyme Reactions
True unimolecular reaction: A single substrate rearranged to a single product
Pseudo unimolecular reaction:
A single substrate molecule is separated into two or more products, normally using water (or other substrate that is in large excess)… kinetically behaves as if it has a single substrate
Bimolecular reactions (involve two substrates):
Sequential bimolecular: Both substrates bind before any product is released
Ordered sequential: Lead substrate binds before the second substrate”
Random sequential: The order of binding does not matter
Non-sequential (ping pong):
The first product needs to be released before the second substrate binds
- Catalytic Mechanisms
General acid & base catalysis:
“Involves groups that donate or accept protons (H⁺), without forming a covalent bond”
Covalent catalysis:
“Involves nucleophilic attachment (electron donation), results in a transient covalent bond”
Metal ion catalysis:
“Ions in the active site introduce ionic interactions”
