Enzymes Flashcards
Briefly describe the effect of an enzyme on a reaction
The activation energy is lowered in an enzyme-catalysed reaction. Thus, more reactant molecules can surmount the energy barrier to reach the transition state to be converted into product molecules
**The total energy difference/free energy change or Gibbs free energy between the reactant molecules and product molecules remains the same.
Briefly describe the four categories of amino acid residues in an enzymes
- Catalytic amino acid residues
- The R groups of these amino acids are directly involved in the catalytic activity, ie. making or breaking of chemical bonds once substrate is bound. - Binding amino acids residues
- The R groups of these amino acids hold the substrate(s) in position via non-covalent bonds while catalysis takes place. - Structural amino acid residues
- Involved in maintaining the specific 3D conformation of the active site, as well as the enzyme as a whole. - Non-essential amino acid residues
- Have no specific functions, and can be removed or replaced without the loss of the enzyme’s catalytic function.
Name the three main types of cofactors:
Inorganic metal ions
Coenzymes
Prosthetic group
Describe the characteristic and purpose of an inorganic metal ion cofactor in an enzyme
Mostly small divalent ions eg. Ca2+
May either be component of active site or affect enzyme activity through allosteric regulation.
Allosteric enzymes have multiple subunits and through conformational changes, bind activators of inhibitors at sites other than the active site. Aforementioned inhibitors usually bind reversibly to the enzyme and act by altering the enzyme’s active and/or allosteric sites to facilitate the catalytic reaction carried out by the enzyme.
Eg. salivary amylase activity is increased in the presence of chloride ions.
Describe the characteristic and purpose of an coenzyme cofactor in an enzyme
Loosely associates with the enzyme during the reaction. Coenzymes act as transient carriers of specific functional groups, hydrogen or electrons. Most coenzymes are derived from vitamins.
Eg. Nicotinamide adenine dinucleotide (NAD) is an important coenzyme in respiration
Describe the characteristic and purpose of an prosthetic group cofactor in an enzyme
Prosthetic groups are tightly bound to the enzyme on a permanent basis.
Eg. the prosthetic group of enzyme catalase is an iron-containing haem group
What is a complete, catalytically active enzyme together with its bound coenzyme and/or metal ions called?
holoenzyme
What is a holoenzyme?
A complete, catalytically active enzyme together with its bound coenzyme and/or metal ions
What is an apoenzyme called?
It is the protein part of such an enzyme
Name the classes of enzymes according to the types of reactions they catalyse (not impt?)
Oxidoreductase - Transfer of electrons (hydride ions or H atoms) aka oxidation-reduction reactions
Transferases - Transfer of functional groups
Hydrolases - Hydrolysis reactions (transfer of functional groups to water)
Lyases - Addition of groups to double bonds, or formation of double bonds by removal of groups
lsomerases - Transfer of groups within molecules to yield isomeric forms
Ligases - Formation of C-C, C-S, C-O, and C-N bonds by condensation reactions coupled to ATP cleavage
Describe how enzymes lowers activation energy
Enzymes lower the activation energy. They do so by:
- orientating the substrates in close proximity, in the correct orientation, to undergo chemical reactions.
- straining critical bonds in the substrate molecule(s), allowing the substrates to attain their unstable transition state.
- providing a microenvironment that favours the reaction (eg. the presence of specific amino acids/ions at the active site may result in a specific set of molecular conditions that favours the formation I breakage of particular bonds).
Explain enzyme specificity using the lock-and-key hypotheses
It suggested that there is an exact fit/complementary shape or conformation between the substrate and the active site of the enzyme, in the same way that a key fits into a lock very precisely.
The enzyme is viewed as a rigid structure, where only substrates that are exactly complementary to the conformation of the active site are able to bind to the active site for catalysis.
Thus, this explains substrate specificity of enzymes.
Explain enzyme specificity using the induced-fir hypotheses
In the induced-fit hypotheses, the enzyme possesses active site flexibility.
The active site does not have a rigid conformation that fits only one type of substrate.
As such, it is rather flexible in conformation and can allow more than one type of substrate to bind.
Moreover, it is not in the precise complementary conformation to the substrate before binding to the substrate.
Upon binding of substrate, the active site changes its conformation slightly to bind the substrate even more firmly/snugly so that the R groups of the catalytic amino acids at the active site are:
- moulded into a specific conformation
- brought into close proximity to the chemical bonds in the substrate hence facilitating catalysis where the substrate is converted to product
Explain the differences between the lock-and-key hypotheses and induced-fit hypotheses for enzyme specificity
The lock-and-key hypotheses explains substrate specificity while the induced fit hypothesis further explains group specificity. Where one enzyme is able to catalyse reactions for a variety of substrates that share similar structural or chemical properties.
In the lock-and-key hypotheses, the enzyme is viewed as a rigid structure, where only substrates that are exactly complementary to the conformation of the active site are able to bind to the active site for catalysis.
In the induced-fit hypotheses, there is active site flexibility, such that the active site does not have a rigid conformation that fits only one type of substrate. Instead, it is rather flexible in conformation and can allow more than one type of substrate to bind
Lastly, in the induced-fit hypotheses, it is not in precise complementary conformation to the substrate before binding to the substrate and changes its conformation slightly to bind to the substrate.
Name the different methods of recording rate of an enzyme-catalysed reaction
Measuring rate of enzyme formation
Measuring initial rate of reaction
Measuring rate of substrate usage
Briefly name all the factors affecting an enzyme reaction
Substrate concentration
Enzyme concentration
Temperature
pH
Explain the effect on low substrate concentration on rate of enzyme-catalysed reaction
Increase in substrate concentration results in a proportional increase in the rate of reaction (substrate concentration is limiting factor).
Not all the active sites of the enzymes are occupied. Rate is limited by the concentration of substrate. An increase in substrate concentration increases the frequency of effective collisions between the enzyme active site and substrate molecules, hence increasing the number of enzyme-substrate complexes formed per unit time and consequently the amount of product formed per unit time, resulting in a proportional increase in the rate of reaction.
Explain the effect of high substrate concentration on rate of enzyme-catalysed reaction
A point will be reached when any further increase in substrate concentration does not result in an increase in the rate of reaction (i.e. the graph reaches a plateau).
The active site of every enzyme molecule is occupied at any given moment. Rate is limited by saturation of enzyme active sites. ie. substrate availability is no longer the limiting factor.
Enzyme concentration is limiting. Any added substrates have to ‘wait’ until existing E-S complexes dissociate to release their products and free enzyme molecules in order to form new E-S complexes.
Further increase in substrate concentration does not result in an increase in the rate of reaction ie. graph reaches a plateau. Substrate concentration is no longer the limiting factor. Rate of reaction can only increase with the addition of enzyme.
Explain the effect of low enzyme concentration on rate of enzyme-catalysed reaction
An increase in enzyme concentration results in a proportional increase in the rate of reaction as the concentration of enzyme is the limiting factor.
The increase in enzyme concentration provides more active sites, and therefore increasing the frequency of effective collisions between substrates and active sites. More enzyme-substrate complexes formed per unit time resulting in an increase in the amount of product formed per unit time. Hence, resulting in an increase in rate of reaction.
Explain the effect of high enzyme concentration on rate of enzyme-catalysed reaction
A point will be reached when any further increase in enzyme concentration does not result in an increase in the rate of reaction ie. the graph reaches a plateau.
Enzyme concentration is no longer a limiting factor. There are not enough substrate molecules competing for the active sites available. Substrate concentration is limiting. Rate of reaction can be increased with the addition of substrate
Explain the effect of temperature increasing on rate of enzyme-catalysed reaction
As the temperature increases to the optimum temperature from 5°C to 40°C, The rate of reaction increases.
At low temperatures near or below freezing point, enzymes are inactivated. Increasing temperature increase the kinetic energy of the substrate and enzyme molecules, thereby increasing the frequency of effective collisions between substrates and active sites, increasing the formation of enzyme-substrate complexes per unit time and the amount of product formed per unit time. This increases the rate of reaction.
Enzyme activity is highest at its optimum temperature of 40°C for most cases. The rate of ES complexes formation is the highest at optimum temperature
If the temperature is increased beyond the optimum temperature of 40°C, a decrease in the rate of reaction occurs despite the increasing frequency of collisions.
Thermal agitation of enzyme molecule disrupts the hydrogen bonds, ionic bonds and other non-covalent interactions that stabilise the specific 3D conformation of the protein molecule. This results in the loss of the 3D conformation of the enzyme and that of its active site so that there is no longer a complementary fit with the substrate. The enzyme is said to be denatured and loses its catalytic function.
The frequency of effective collisions between substrates and active sites decrease and the rate of formation of enzyme-substrate complexes drops. Hence, less product is formed per unit time.
Explain the effect of deviation in pHon rate of enzyme-catalysed reaction
Changes in pH can affect enzyme activity by altering the ionic charge of the acidic and basic R groups of proteins:
- Lower pH»_space; more H+ ions available to neutralise negative charges present in the enzyme.
- Higher pH»_space; less H+ ions available to neutralise negative charges present in the enzyme.
Change in ionisation of amino acids disrupts the ionic bonds/hydrogen bonds that maintain the 3D conformation of the enzyme, denaturing the enzyme.
Briefly explain the rationale behind an optimum pH
At the optimum pH, the rate of reaction is at a maximum .
Intra-molecular bonds are intact and conformation of active site is ideal for binding, therefore frequency of effective collision is the highest with largest amount of enzyme-substrate complexes formed.
What is the Michaelis Constant?
Km is measured as the substrate concentration that allows an enzyme-catalysed reaction to proceed at half the maximum velocity, 1/2Vmax
