Enzymes- Introduction to enzymes, regulation of enzyme activity and enzymes in diagnosis, prognosis and therapy Flashcards
What are enzymes and what do they do?
- Biological catalysts of chemical reactions
- Specific action on particular biochemical compounds (substrates)
- Nearly all are proteins
- Increase the rate at which the reaction equilibrium is reached, but do NOT shift the position of equilibrium
Activation energy
Difference of free energy
Transition state
The highest energy
How do enzymes decrease EA?
- by providing catalytically competent groups for a specific reaction mechanism
- by binding substrates such that their orientation is optimised for the reaction
- by preferentially binding and stabilising the transition state(s) of the reaction
What is the AS of an enzyme
- is the region of the enzyme at which substrate binding and conversion to product takes place;
- is a 3-dimensional space comprising crucial amino acid residues;
- may represent only a small part of the protein structure;
- binds substrate via multiple weak interactions;
- provides substrate specificity because of its unique 3D arrangement of atoms
How is enzyme activity measured?
Use the Michaelis- Menten model
Km
Substrate concentration at which the rate is half Vmax
Vmax
Maximum rate of reaction
Factors affecting enzyme-catalysed reaction rates
- Substrate/enzyme concentration
- Temperature
- pH
- Inhibitors, both natural and exogenous
Irreversible enzyme inhibition
Covalent modification of the enzyme, usually at amino acid side chains in the AS
Competitive inhibition and how Km is affected
enzyme
• Inhibitor usually binds same site as substrate (active site)
• Inhibition can be overcome by high substrate concentration
• In the plot for double reciprocal, the lines have the same Vmax because the inhibitor gets swamped by the substrate concentration. However, Km is increased – more substrate is needed to reach half of Vmax.
Non-competitive inhibition and how Km is affected
- Inhibitor and substrate can bind simultaneously at independent sites
- Inhibitor alters conformation or accessibility of active site
- Inhibition not affected by high substrate concentration
- Vmax decreases (some of the enzyme unusable), Km is unchanged , increasing substrate concentrate has no effect.
Non-steroid anti-inflammatory drug examples and what they do
Aspirin:
• Covalent modification of a serine residue in the active site
• Inhibitor binding is competitive, inhibition is irreversible
Ibuprofen:
• Binds to active site, but not covalently attached
• Inhibitor binding is competitive, inhibition is reversible
IC50
The inhibitor concentration at which 50% of activity remains for a standard amount of enzyme and substrate
Cofactors are essential for enzyme function- metal ions
- Includes major minerals such as Mg2+ and Ca2+ and trace elements such as Cu2+, Zn2+, Fe2+/3+, Mn2+ and Mo4+
- Metal ions can be part of active site and/or be involved in electrostatic substrate binding (Zn2+ in carbonic anhydrase, Mg2+ in kinases)
- Metal ions may act as redox agents (Fe2+/3+ in cytochrome p450s)
- Metal ions may regulate activity of enzymes (Ca2+ in calpain)
Cofactors are essential for enzyme function- coenzymes
• Coenzymes function as carriers of reaction components:
- NADH and FADH2 carry electrons (‘reducing power’)
- Coenzyme A carries acyl units
- Biotin and thiamine pyrophosphate carry CO2 units (bound to carboxylases)
Effect of mutations in enzymes
Glucose-6-phosphate dehydrogenase (G6DPH) deficiency causes metabolic defects
- X-linked recessive; most carriers asymptomatic (disease without showing symptoms)
- The enzyme produces a large proportion of the body’s NADPH needed to drive biosynthesis of nucleic acids, lipids etc.
- Where symptomatic, symptoms can include haemolytic crises (accelerated rate of blood destruction), jaundice (can lead to brain damage (kernicterus) in infants)
Favism
A disease that results from a deficiency of the enzyme glucose-6-phosphate dehydrogenase (G6PD). Favism usually is due to a genetic disorder.
