Enzyme properties Flashcards
Enzymes definition
-Biological catalysts that speed up rate of reaction,
-Without altering the final equilibrium between reactants and products
How do enzymes affect the activation energy?
-They lower the activation energy of a reaction
What is the induced fit theory?
-Enzymes undergo conformational changes upon substrate binding
-Specific functional groups within the enzyme are brought into proper position to catalyse the reaction
What is a transition state and how does enzyme use it to speed up reaction?
-Transition state is an unstable, high energy intermediate in a reaction
-Enzyme is complementary to the transition state
What is substrate specificity?
Enzymes will:
-Catalyse only 1 type of reaction
-Act on only a few related molecules
-Act only on one isomer if two stereoisomer forms exist
What are the 7 enzyme classes?
-Oxidoreductases
-Transferases
-Hydrolases
-Lyases
-Isomerases
-Ligases
-Translocases
Why are enzymes very sensitive to changes in temperature?
-Enzyme structure is stabilised by many weak bonds (hydrogen, van der waals, hydrophobic)
-Weak bonds are easily broken causing a disorganised structure with no catalytic activity
How does pH affect enzyme-catalysed reactions?
-Unfolding of enzyme leading to complete inactivation
-Changes in ionisation state of active site residues leading to change in Km and Vmax
-Changes in ionisation state of the substrate leading to additional acid/base catalysis
-Changes leading to altered binding to the enzyme and hence a change in Km
What is enzyme kinetics?
Study of the rate of an enzyme catalysed reaction
-How that rate varies with different substrate concentrations
-Effects of inhibitors
What are hyperbolic kinetics?
-At low substrate concentration, reaction rate is directly proportional to substrate concentration (first order reaction)
-At high substrate concentration, the reaction rate is independent of the substrate concentration (zero order reaction)
What are 3 assumptions in the Michaelis-Menten equation?
-More substrate than enzyme so amount of substrate bound by enzyme at any one time is small
-Initial velocities are measured, so the concentration of product is small and the reverse reaction can be ignored
-Enzyme Substrate complex is in a steady state, does not change with time. Formation of ES = Breakdown of ES
What does the Michaelis-Menten equation mean?
What is the definition of Km?
-The substrate concentration we need so that the enzyme is functioning at half its maximum velocity
When does Km truly represent the affinity of an enzyme for its substrate in the ES complex
-When the enzyme substrate complex is formed, it dissociates more rapidly than the product is formed
-Then Km = dissociation constant of the ES complex
-k2 is ignored
What is the Kcat- turnover number
-Number of substrate molecules converted to product in a unit of time on a single enzyme molecule
-when the enzyme is saturated with substrate
What is the best method for comparing catalytic efficiency?
-Kcat/Km, the specificity constant, provides the best method
-2 enzymes may have the same Kcat but very different Km
Advantages of linearising a hyperbole (Lineweaver-Burk double reciprocal plot)
-Reduces the number of experiments needed (2 for a straight line)
-Not depending on solubility of substrate and products. Can take several measurements at whatever concentration
How do competitive inhibitors affect the Km and Vmax?
-Increase apparent Km
-Do not alter Vmax
How do competitive inhibitors block the enzyme active site
-Interfere with first step (recognition between enzyme and substrate) but no second step
How do non-competitive inhibitors affect Km and Vmax?
-Do not alter Km because they do not interfere with recognition between enzyme and substrate
-Decrease Vmax
How do non-competitive inhibitors interfere with the catalytic mechanism?
-Interferes with the second step
-Once bound, the enzyme can’t use electron transport to carry out reaction
What are irreversible enzyme inhibitors
-They kill catalytic activity of an enzyme
-Examples are insecticides and nerve agents
-AChE (Acetylcholinesterase) inhibitors
-Do not follow Michaelis-Menten behaviour because reaction is irreversible
What enzymes have a sigmoidal shape curve?
-Allosteric enzymes
How is sigmoidal response in allosteric enzymes created?
-Co-operative substrate binding
-Sigmoidal response can be considered to be a result of combining 2 Michaelis-Menten enzymes:
-One with a high value of Km that corresponds to Tense state
-One with a low value for Km that corresponds to relaxed state
How do allosteric inhibitors and activators affect the reaction?
-Allosteric inhibitors require more substrate and favours the tense state
-Allosteric activators require less substrate and favours the relaxed state
What is feedback inhibition of metabolic pathways?
-End product of a pathway can affect the beginning of a pathway to control rate of reaction
-Allosteric enzymes allow feedback inhibition
Explain regulation by covalent modification
-Reversible addition or removal of phosphate from amino acid residues by kinase and phosphatase regulatory enzymes
-Phosphorylated enzyme may be active
-Phosphorylated enzyme may be inactive
How does insulin regulate blood sugar?
-It increases the rate of synthesis of key enzymes involved in glucose metabolism: Glucokinase, Phosphofructokinase, Pyruvate kinase
Summarise some of the important mechanisms for regulation of enzyme activity
