WJEC AS Biology Unit 1.4 - Enzymes and Biological Reactions

Metabolism

• Series of enzyme controlled reactions

• Combination of anabolic and catabolic reactions that are catalysed by enzymes

Anabolic reactions

Reactions which construct molecules from smaller units, eg protein synthesis

Catabolic reactions

Reactions when break down molecules into smaller units, eg digestion

Enzyme

• Biological catalysts

• Speed up reactions without being changed themselves

• Found in living things

• All are proteins

• Often end in -are, e.g. amylase

Catalyst

An arm or molecule that alters the rate of a chemical reaction without taking part in the reaction or being changed by it

Substrate

The substance on which an enzyme acts

Enzyme-substrate complex

• Intermediate structure formed during an enzyme-catalysed reaction in which the substrate and enzyme bind temporarily, such that the substrates are close enough to react

• Forms a product

Product

The substances that are made, e.g. maltose etc.

Active site

The specific three-dimensional site on an enzyme molecule to which the substrate binds by weak chemical bonds. Formed by amino acids

Specific

Only one type of substrate can bind

Denaturation

• The breakdown of the bonds that hold the protein together, including the active site, in shape

• This changes the 3D structure of the enzyme incl the active site so the substrate can no longer bind

Activation energy

The minimum energy that must be put into a chemical system for a reaction to occur

Collision theory of enzyme reactions

For molecules to react, they have to collide with energy to break and form bonds

Lock and key model

• The specific, unique shape of the active site means that an enzyme can only catalyse one type of reaction

Induced fit model

Where changed of shape of both active site and substrate being reactive groups of enzyme and substrate close to each other, weakening bonds in the substrate so the reaction takes places at a lower activation energy

Sites of enzyme action

• Extracellular (inside cells); some enzymes are secreted from cells by exocytosis and catalyse extracellular reactions

• Intracellular (outside cells);

  • In solution; intracellular enzymes may act in solution inside cells

  • Membrane-bound; intracellular enzymes may be attached to membranes such as on the crustal of mitochondria

Catalysis

The lowering of activation energy

Metabolic pathway

• A sequence of enzyme-controlled reaction in which a product of one reaction is a reactant in the next

• Many enzymes can be close together and can catalyse separate reactions in a series

Structure of enzymes

• Globular proteins

• Proteins with tertiary structure

Properties of enzymes

• Speed up reactions

• Not used up

• Not changed

• Catalyse many reactions per second

• Only catalyse reactions that are energetically favourable and would happen anyway

Protein nature of enzymes

• Globular proteins

• Proteins with tertiary structure

• The protein chain folds into a spherical or globular shape with hydrophilic R groups on the outside of the molecule, —> enzymes = soluble

• Each enzyme has a particular sequence of amino acids

• The elements in the R groups determine the bonds the amino acids make with each other

• These bonds (eg hydrogen bonds, ionic bonds, disulphide bridges) hold the enzyme molecules in its tertiary form

• Small area with specific 3D shape = active site which gives enzyme many of its properties

Effect of temperature on enzyme controlled reactions

• Increased temp —> increased KE of enzyme + substrate molecule; collide with enough energy more often, increasing rate of reaction

• Above optimum, molecules have more KE but rate goes down as increasing vibration breaks H bonds, changing tertiary structure

• Alters shape of active site and substrate will not fit. Enzyme = denatured, a permanent change in structure

• Low temps, enzyme = inactivated as molecules have v low KE. Shape = unchanged and enzyme will work again if the temp is raised

Effect of pH on enzyme controlled reactions

• At optimum, rate = highest

• Small changes around the optimum cause small reversible changes in enzyme structure and reduce its activity, but extremes of pH denatured enzymes

• Changes on amino acid side chains of active site age affected by hydrogen ions or hydroxide ions

• Low pH, excess H ions = attracted to - charges + neutralise them

• High pH, excess OH ions neutralise + charges

• Disrupts ionic and H bonds maintaining shape of active site

• Shale changes, denaturing the enzyme

• No e-s complexes form + enzyme activity is lost

Effect of enzyme concentration when substrate is unlimited

• product leaves active site, enzyme molecule can be reused, so only low enzyme conc is needed to catalyse a large number of reactions

• Number of substrate molecules that one enzyme molecule can turn into products in a given time is the turn-over number

• As the enzyme conc increases, there are more active site available + for increases

Effect of substrate concentration when enzyme concentration is limited

• if enzyme conc is constant, rate increases as substrate conc increases

• At low substrate conc the enzyme molecules have only a few substrate molecules to collide with. Active sites are not colliding to full capacity

• With more substrate, more active sites filled

• Conc of substrate is controlling the ror and so is a limiting factor

• More substrate is added, at a critical conc, all the active sites become occupied and rate is at its max

• Active sites all full, enzyme is saturated

Effect of competitive inhibitors

• the greater the substrate conc, in relation to inhibitor, the greater the chance that the substrate and enzyme will collide

• No reaction takes place to form products

• Effect is overcome when the substrate is in excess

Effect of non-competitive inhibitors

• as inhibitor conc increases, the rate of reaction and final mass of product decrease

Enzymes and activation energy

• Modify the substrate so the reaction requires a lower activation energy

Inhibitor

A molecule or ion that binds to an enzyme and reduces the rate of reaction the enzyme catalyses. Can be reversible or irreversible

Competitive inhibition

• Reduction of the rate of an enzyme-controlled reaction by a molecule or ion that has a complementary shape to the active site, similar to the substrate, and binds to the active site, preventing the substrate from binding

• Have a molecular shape complementary to the active site and similar to that of the substrate

• Fits in enzyme active site —> e-I complex, blocking active site

• Prevents binding of substrate

• Inhibitor and substrate compete for active site

• Substrate molecules unable to occupy active site

• No reaction takes place to form products

Non-competitive inhibition

• An atom, molecule or ion that reduces the rate of an enzyme-controlled reaction by binding to the enzyme at a position other than the active site, altering the shape of the active site and preventing the substrate from successfully binding to it

• Bind to enzyme somewhere other than active site - an allosteric site

• Substrate and inhibitor do not compete with the substrate

• Affect bonds within enzyme molecule and alter its overall shape, including that of the active site, distorts shape of active site

• Substrate cannot bind with the active site, and no enzyme-substrate complexes form

• If substrate conc is increased, the degree of inhibition is unaffected

• Rate of reaction is always lower when nci is present

End-point inhibition

• product of enzyme-controlled reaction

• Acts as an inhibitor

• Binds to enzyme, slowing down its own production using negative feedback

Immobilised enzymes

• Enzyme molecules bound to an inert material, over which the substrate molecules move

• Can be packed into glass columns

• Substrate is added to the top of the column and as it flows down, its molecules bind to the enzyme molecules’ active sites, both on the bead surface and inside the beads as the substrate molecules diffuse in

• Column can be used repeatedly

• Enzyme is fixed and does not contaminate products, therefore easy to purify

Surface area of immobilised enzymes

• Large beads —> smaller total surface area then if the same volume had been used to make small beads

• Smaller beads; substrate molecules will have easier access to enzyme molecules and so they will produce a higher rate of reaction

Effect of immobilisation of enzymes on rate of reaction

• Makes enzymes more stable as it creates a micro environment allowing reactions to occur at higher temperatures or more extreme pHs than normal

• Prevents shape change that would denature its active side so the enzyme can be used in a wider range of physics conditions

• Enzymes immobilised in beads = lower rate of reaction than those immobilised on a membrane, if all other factors are constant

• Some of the active sites are inside the beads + the substrate takes time to diffuse them

• On a membrane = readily available for binding, so give a higher rate of reaction

Benefits of immobilised enzymes overall

• Increased stability and function over a wider range of environments that enzymes free in solution

• Products are not contaminated with the enzyme

• Enzymes are easily recovered for reuse

• A sequence of columns can be used so several enzymes with differing pH or temp optima can be used in one process

• Enzymes can be easily added or removed, giving greater control over the reaction

Benefits of immobilised enzymes in industry

• Separated easily from products and reused

• More thermo- and pH-stable

• Several enzymes can be used in the same enzyme continuous fermenters (reactors) despite having different optimum condition

• Products can be removed continuously

• Products are pure and free of enzyme

Enzymes in biosensors

• Turn a chemical signal into an electrical signal

• Rapidly + accurately detect, identify and measure even very low concentrations of important molecules - high sensitivity

• Enzymes = specific and are able to select one type of molecule from a mixture, even at very low concentrations

• Example; detection of blood glucose

• Enzymes can be immobilised onto test strips, where different strips may detect a variety of molecules

Uses of immobilised enzymes

• Lactose-free milk; milk is passed down a column containing immobilised lactase. The lactose binds to the active sites on the lactase and is hydrolysed to its components

• Biosensors

• High-fructose corn syrup (HFCS) manufacture