chapter 2: enzymes! Flashcards
what are the general properties of enzymes?
- enzymes are highly specific
- an enzyme with absolute specificity only catalyses a single specific reaction in a single specific substrate
- an enzyme with group specificity catalyses a single specific reaction in a group of related substrates - enzymes are highly efficient in very small amounts
- enzymes have a high turnover number ( number of substrate molecules converted into products per unit time by one molecule of enzyme)
- this is because the enzyme remains unchanged at the end of th reaction, and the active site can be used repeatedly - enzymes catalyse reversible reactions
- enzymes speed up rate of both forward and backward reactions until equilibrium is reached in a reversible reaction
> enzymes speed up the rate at which equilibrium is reached
- enzymes do not alter position of equilibrium
- the presence of enzymes also does not alter the properties of the end-products of the reaction they catalyse - enzymes are affected by reaction conditions
- can be affected by various factors like enzyme concentration, substrate concentration, temperature, pH and inhibitors
mode of enzyme action
what is the energy profile of a reaction?
- a chemical reaction involves both bond breaking and bond forming
- it requires energy to contort reactant molecules into a highly unstable state before bonds can be broken and new ones can be formed
- the initial investment of energy to contort reactant molecules so the bonds can break is known as free energy of activation, or activation energy (Ea)
- Ea is the energy barrier that has to be overcome before a reaction can occur and it determines the rate of reaction
> higher activation energy will lead to a slower rate of reaction - Ea is often supplied in the form of thermal energy (heat) absorbed by the reactant molecules from the surroundings
absorption of thermal energy can lead to:
- increase in speed of reactant molecules and thus an increase in frequency of collisions between reactant molecules
- thermal agitation of atoms within molecules, causing bonds to break more easily
- when the reactant molecules absorb sufficient energy for bonds to break, the reactants are known to reach an unstable condition known as the transition state
- once bonds are broken, atoms will settle into their new and more stable bonding arrangements and energy is released into the surrounding
- an enzyme works by lowering the activation energy of the reaction it catalyses
- increasing the rate of reaction and the rate at which equilibrium is reached
mode of enzyme action
describe the enzyme structure and active site
- an enzyme consists of four categories of amino acids
- catalytic residues: make or break chemical bonds; responsible for the enzyme’s catalytic activity
- contact/ binding residues: hold substrate(s) in place during catalysis, by forming temporary interactions like weak hydrogen bonds with the substrate
-** structural residues:** maintain the correct specific 3D conformation of the active site so that enzyme can function properly
- non-essential residues: near the surface of enzyme; no specific functions thus can be removed or replaced without loss of function - the active site of an enzyme is a small pocket/ groove/ cleft on its surface and is usually formed by only a few (3 to 12) amino acid residues
- it has a specific 3D conformation which is complementary to that of its substrate
- the active site contains catalytic and binding residues and the site of catalysis
what are the three steps in the enzyme’s mode of action?
step 1: formation of enzyme-substrate complex
- both the specific 3D conformation and charges present at the active site allow only specific substrates with a 3D conformation and charges that are complementary to the active site to enter it in a specific orientation
- a small rearrangement (induced fit) of chemical groups may occur in both the enzyme active site and the substrate molecules
- certain binding residues of the active site may form weak, temporary but extensive bonds with substrates, forming the enzyme-substrate complex
- weak bonds are usually hydorgen bonds
step 2: catalysis / lowering of activation energy
- interactions between the enzyme ans substate molecules occur, lowering tthe activation enerfy and increasing the rate of the enzyme-catalysed reaction
lowering the activation may occur by:
- bringing substrates close together with each other at the active site
- bringing substrates into the correct orientation with each other to react
- altering the charges on the substrates
- physically straining bonds within substrates
- creating a more conducicen micro-environment for substates to react
step 3: product formation and regeneration of enzymes
- when catalysis is completed, the product(s) are no longer complementary in 3D conformation and charge to the active site and thus leave the active site
- the temporary changes in 3D conformation and charge at the active site are reversed and the active site is now availible again for substrate binding
-
enzyme specificity
how does the ‘lock and key’ hypothesis explain why enzymes are specific?
- the enzyme’s (lock) active site has a fixed 3D conformation produced by the folding of the polypeptide chain
- this fixed 3D conformation is perfectly complementary in shape to the substrate (key)
enzymes specificity
how does the induced fit hypothesis explain enzyme speecificity?
- this hypothesis states that the active site is physically flexible
- the active site is able to mould itself around the substrate
- an enzyme thus not a still structure that is of fixed 3D conformation
- > the active site is also not a rigid groove to hold the substrate
- the 3D conformation of the active site of the enzyme molecule is initially not complementary to the 3D conformation of substrate
- the substrate entering the active site induces a slight change in the active site’s 3D conformation due to interactions between the substrate’s chemical groups and the R groups of the amino acid residues that form the active site
- the 3D conformation change causes the active site to fit more snugly around the substrate
- the shape changes in the active site to fit the substrate molecule better is called induced sit
- induced fit brings catalytic residues into positions that improve their ability to catalyse the chemical reaction
- after products are formed, they no longer fit into the active site and leaves the active site
> ( no longer complementary in 3D conformation and charge)
> the enzyme molecule reverts to its original conformation and it is free to bind another substrate molecule
rate of enzyme-catalysed reactions
how can we measure the rate of enzyme-catalysed reactions?
- can be measure by the amount of substrate depleted or used over time
- or the amount of products formed over time
- the actual rate of reaction can be calculated by measuring the slope of the tangent at the initial part of the curve
- the steeper the slope, the higher the rate
rate of enzyme-catalysed reactions
what are some factors affecting the rate of enzyme-catalysed reactions?
-enzyme concentration, substrate concentration, temperature and pH
> when considering one factor, other factors must be kept constant
factors affecting the rate of enzyme-catalysed reactions
how does enzyme concentration affect the rate of enzyme-catalysed reactions?
- when the enzyme concentration is increased, the rate of reaction increases proportionally to the increase in enzyme concentration
- this is because there are more active sites available for substrates to bind at
> leading to an increase in the frequency of effective collisions between enzymes and substrates
> increase in the rate of enzyme-substrate complex formation and
> the increase in the rate of product formation - provided that temperature and other physical conditions are kept constant during the reaction and the substrates are in excess
> the rate of reaction is directly proportional to enzyme concentration - if the substrate concentration is limiting, further addition of enzyme will not lead to an increase in the reaction rate
> there are many more empty active sites that there are substrates availible
factors affecting the rate of enzyme-catalysed reactions
how does substrate concentration affect the rate of enzyme-catalysed reactions?
- the reaction for most enzyme-controlled reactions varies witht he availibility of substrate
- when the enzyme concentration is in excess, an increase in the substrate concentration will lead to an increase in the rate of reaction
- at low substate concentrations, the rate of the reaction increases with increasing substrate concentrations
> many enzyme molecules have availible active sites which are unoccupied and the limited number of substrate molecules largely determines the reaction rate
> substrate concentration is the limiting factor - increasing substrate concentration will increase the reaction rate as more active sites are being occupied and more enzyme substrate complexes and products are formed per unit time
- beyong a certain high substrate concentration, when there are too many substrate molecules, the rate of reaction will reamin constant as all the active sites are being occupied by substrate molecules all at once
- all the active sites are saturated
- this will be the theoretical maximum rate of reaction for that enzyme and is reffered to as its Vmax
- beyond this substrate concentration and the rate of reaction is still constant,
> an increase in rate can only be achieved by increasing enzyme concentration
> substrate concentration is thus no longer the limiting factor, with enzyme concentration now being the limiting factor
how does temperature affect the rate of reaction?
(temperature coefficient)
how does temperature affect the rate of reaction?
(below optimum temperature)
- at very low temperatures, the enzymes are inactive
- rate of reaction is very low as the frequency of effective collisions between substrate and enzyme molecules is low
- as there is an increase in temperature, both the enzyme and substrate molecules gain kinetic energy
- there is an increase in frequency of effective collisions between substrate and enzyme molecules
- more ES complexes are formed per unit time and therefore more products are formed per unit time
- rate of reaction increases until optimum temperature is reached
how does temperature affect the rate of reaction?
(at optimum temperature)
- rate of reaction is fastest as there is greatest freqeuncy of effective collsions between enzymes and substate molecules
- rate of ES complex formation is the highest
how does temperature affect the rate of reaction?
(above optimum temperature)
- rate of reaction decreases despite an increase in the frequency of collisions
- increasing temperature increases thermal agitation of enzyme molecules
- this casues R group interactions like hydrogen bonds, ionic bonds and hydrophobic interactions in tertiary and quaternary structures to be disrupted
- hydrogen bonds in the secondary structures may also be disrupted
- enzyme molecules are denatured as the precise 3D conformation of the active site is changed
> so no longer complementary in 3D conformation to the substrate and cannot bind the substrate - catalytic activity of the enzyme is lost, leading to a lower rate of ES complez formation and thus lower rate of reaction ( product formation)
- disulfide bonds are strong covalent bonds are usually broken only at very high temperatures by high heat energy but protein denaturation usually occurs before it can happen
