This power-point presentation will give a complete overview about enzymes, nomenclature of enzymes. Enzymes inhibition is also covered in this ppt. Along with some basin introduction to G- protein coupled receptors is also provided.
3. Introduction to Enzymes
Enzymes are biocatalysts - the catalysts of life.
A catalyst is defined as a substance that increases the velocity or rate of a chemical
reaction without itself undergoing any change in the overall process.
Enzymes may be defined as biocatalysts synthesized by living cells. They are protein in
nature (exception - RNA acting as ribozyme), colloidal and thermolabile in character, and
specific in their action.
For example, In the laboratory, hydrolysis of proteins by a strong acid at 100'C takes at
least a couple of days. The same protein is fully digested by the enzymes in
gastrointestinal tract at body temperature (37'C) within a couple of hours. This
remarkable difference in the chemical reactions taking place in the living system is
exclusively due to enzymes.
History:
• Berzelius in 1836 coined the term catalysis “Greek: to dissolve”.
• In 1878, Kuhne used the word enzyme “Greek: in yeast”.
• Isolation of enzyme system from cell-free extract of yeast was achieved in 1883 by
Buchner.
• Sumner first achieved the isolation and crystallization of the enzyme urease from jack
bean and identified it as a protein.
5. Previously, enzymes were given by their discoverers in an arbitrary
manner. Sometimes, the suffix-ase was added to the substrate for
naming the enzymes e.g. lipase acts on lipids. These are known as
trivial names of the enzymes which, however, fail to give complete
information of enzyme reaction.
The International Union of Biochemistry (lUB) appointed an Enzyme
Commission in 1961. The committee made a thorough study of the
existing enzymes and devised some basic principles for the
classification and nomenclature of enzymes. IUB classified the
enzymes into 6 major classes.
6. 1. Oxidoreductases : Enzymes involved in oxidation-reduction reactions.
2. Transferases :Enzymes that catalyze the transfer of functional groups.
3. Hydrolases : Enzymes that bring about hydrolysis of various
compounds.
4. Lyases : Enzymes specialized in the addition or removal of water,
ammonia, CO2 etc.
5. Isomerases :Enzymes involved in all isomerization reactions.
6. Ligases : Enzymes catalyzing the synthetic: Reactions (Greek : ligate-
to bind) where two molecules are joined together and ATP is used.
7. Chemical nature and properties of
enzymes
The functional unit of the enzyme is known as holoenzyme which is often made up of
apoenzyme (the protein part) and a coenzyme (non-protein organic part)
Holoenzyme Apoenzyme + Coenzyme
(active enzyme) (protein part) (non-protein part)
The term prosthetic group is used when the non-protein moiety tightly (covalently)
binds with the apoenzyme.
The word monomeric enzyme is used if it is made up of a single polypeptide e.g.
ribonuclease, trypsin.
Some of the enzymes which possess more than one polypeptide (subunit) chain are
known as oligomeric enzymes e.g. lactate dehydrogenase, aspartate transcarboxylase
etc.
8. Factors affecting enzyme activity
Concentration of enzyme- As the concentration of the enzyme is increased, the velocity of the
reaction proportionately increases.
Concentration of substrate- Increase in the substrate concentration gradually increases the
velocity of enzyme reaction within the limited range of substrate levels.
Effect of temperature- Velocity of an enzyme reaction increases with increase in temperature
up to a maximum and then declines. A bell-shaped curve is usually observed.
Effect of pH- Increase in the hydrogen ion concentration (pH) considerably influences the
enzyme activity and a bell-shaped curve is normally obtained.
Effect of product concentration- The accumulation of reaction products generally decreases
the enzyme velocity.
Effect of activators- Some of the enzymes require certain inorganic metallic cations like Mg2+,
Mn2+, zn2+, ca2+, co2*, cu2+, Na+, K+ for their optimum activity. Metals function as
activators of enzyme velocity and bring about conformational changes.
Effect of time- Under ideal and optimal conditions (like pH, temperature etc.), the time
required for an enzyme reaction is less.
Effect of light and radiation- Exposure of enzymes to ultraviolet, beta, gamma and X-rays
inactivates certain enzymes due to the formation of peroxides.
10. Theories of mechanism of enzyme action
Lock and Key model
Or
Fischer's template
theory
Induced fit
theory or
Koshland's
model
Substrate
strain theory
12. Enzyme inhibitor is defined as a substance which binds with the
enzyme and brings about a decrease in catalytic activity of that
enzyme.
The inhibitor may be organic or inorganic in nature.
There are three broad categories of enzyme inhibition.
1. Reversible inhibition.
2. Irreversible inhibition.
3. Allosteric inhibition
13. Reversible inhibition
The inhibitor binds non-covalently with enzyme.
The enzyme inhibition can be reversed if the inhibitor is removed.
The reversible inhibition is further sub-divided into
l. Competitive inhibition
ll. Non-competitive inhibition
Competitive Inhibition:
• The inhibitor which closely resembles the real substrate is regarded as a “substrate
analogue”
• The inhibitor competes with substrate and binds at the active site of the enzyme
but does not undergo any catalysis
• Eg. Methanol is toxic to the body when it is converted to formaldehyde by the
enzyme alcohol dehydrogenase (ADH). Ethanol can compete with methanol for
ADH. Thus, ethanol can be used in the treatment of methanol poisoning.
14. • Another example of competitive reversible inhibition: Antimetabolites (5-fluorouracil) are
the chemical compounds that block the metabolic reactions by their inhibitory action on
enzymes. Antimetabolites are usually structural analogues of substrates and thus are
competitive inhibitor.
Non competitive inhibition
• The inhibitor binds at a site other than the active site on the enzyme surface.
• This binding impairs the enzyme function.
• The inhibitor has no structural resemblance with the substrate.
(A) Competitive Inhibition
(B) Non-competitive
inhibition.
15. Irreversible inhibition
The inhibitors bind covalently with the enzymes and inactivate them, which is
irreversible. These inhibitors are usually toxic poisonous substances.
Examples:
• Iodoacetate is an irreversible inhibitor of the enzymes such as papain and
glyceraldehyde 3-phosphate dehydrogenase. Iodoacetate combines with sulfhydryl
(-SH) groups at the active site of these enzymes and makes them inactive.
• Di-isopropyl fluorophosphate (DFP) is a nerve gas developed by the Germans
during Second World War. DFP irreversibly binds with enzymes containing serine at
the active site, e.g. serine proteases, acetylcholine esterase.
• Many organophosphorus insecticides like melathion are toxic to animals (including
man) as they block the activity of acetylcholine esterase, resulting in paralysis of
vital body functions.
16. • Alcohol gets metabolized by aldehyde dehydrogenase. Aldehyde that is
produced gets converted to acetic acid. Disulfiram irreversibly inhibits
enzyme aldehyde dehydrogenase. Thus accumulation of aldehyde makes a
person sick and thus alcohol consumption is stopped.
• The penicillin antibiotics act as irreversible inhibitors of serine - containing
enzymes, and block the bacterial cell wall synthesis.
17. Suicide inhibition:
• Specialized form of irreversible inhibition.
• Original inhibitor (structural analogue or competitive inhibitor) is
converted to more potent form by the same enzyme that is to be
inhibited.
• The so formed inhibitor binds irreversibly with the enzyme.
• A good example of suicide inhibition is allopurinol (used in the treatment
of gout) Allopurinol, an inhibitor of xanthine oxidase, gets converted to
alloxanthin, a more effective inhibitor of this enzyme.
18. Allosteric Inhibition
Allosteric inhibition is the slowing down of enzyme-catalyzed chemical reactions that occur in
cells.
These metabolic processes are responsible for the proper functioning and maintenance of our
bodies’ equilibrium, and allosteric inhibition can help regulate these processes.
Allosteric inhibitors slow down enzymatic activity by deactivating the enzyme.
An allosteric inhibitor is a molecule that binds to the enzyme at an allosteric site. This site is
not at the same location as the active site.
Upon binding with the inhibitor, the enzyme changes its 3D shape.
It is a form of noncompetitive inhibition
Allosteric inhibitors prevent the body from wasting energy to create unnecessary products.
Examples:
• In glycolysis (cellular respiration), enzyme Phosphofructokinase is responsible for conversion of
ADP to ATP. Once there is an excess of ATP in the system, ATP acts as allosteric inhibitor. It
binds to phosphofructokinase to slow down the conversion of ATP. In this way, ATP is
preventing the unnecessary production of itself.
20. They are membrane receptors that are coupled to intracellular effector systems via a
G-protein.
After stimulation of these receptor, action is produced within seconds
They constitute the largest family, and include receptors for many hormones and slow
transmitters.
For example the muscarinic acetylcholine receptor, adrenergic receptors and
chemokine receptors.
21. Molecular Structure
G-protein-coupled receptors consist of a single polypeptide chain of up to 1100 residues.
Their characteristic structure comprises seven transmembrane α helices, similar to those of the
ion channels, with an extracellular N-terminal domain of varying length, and an intracellular C-
terminal domain.
GPCRs are divided into three distinct families.
They share the same seven-helix (heptahelical) structure, but differ in other respects, principally
in the length of the extracellular N terminus and the location of the agonist binding domain.
22. G-Protein
G-proteins comprise a family of membrane-resident proteins whose
function is to recognize activated GPCRs and pass on the message to the
effector systems that generate a cellular response.
They actually called G-proteins because of their interaction with the
guanine nucleotides, GTP and GDP.
Structure :
• G-proteins consist of three subunits: α, β and γ.
• Guanine nucleotides bind to the α subunit, which has enzymic activity,
catalyzing the conversion of GTP to GDP.
• The β and γ subunits remain together as a βγ complex.
• All three subunits are anchored to the membrane through a fatty acid
chain, coupled to the G-protein through a reaction known as prenylation
23. Step involved in action of G protein
In the 'resting' state, the G-protein exists as an unattached αβγ trimer, with GDP occupying
the site on the α subunit.
When a GPCR is activated by an agonist molecule, a conformational change occurs, involving
the cytoplasmic domain of the receptor, causing it to acquire high affinity for αβγ.
Association of αβγ with the receptor causes the bound GDP to dissociate and to be replaced
with GTP (GDP-GTP exchange),
which in turn causes dissociation of the G-protein trimer, releasing α-GTP and βγ subunits;
these are the 'active' forms of
which diffuse in the membrane and can associate with various enzymes and ion channels,
causing activation of the target.
Association of α subunits with target enzymes can cause either activation or inhibition,
depending on which G protein in involved.
the G-protein, Signaling is terminated when the hydrolysis of GTP to GDP occurs through the
GTPase activity of the α subunit.
The resulting α-GDP then dissociates from the effector, and reunites with βγ, completing the
cycle.
Four main classes of G-protein (Gs, Gi, Go and Gq) are of pharmacological importance.
24.
25. Targets for G-Proteins
The main targets for G-proteins, through which GPCRs control
different aspects of cell function
• Adenylyl cyclase, the enzyme responsible for cAMP formation
• Phospholipase C, the enzyme responsible for inositol phosphate
and diacylglycerol (DAG) formation
• Ion channels, particularly calcium and potassium channels
• Rho A/Rho kinase, a system that controls the activity of many
signaling pathways controlling cell growth and proliferation,
smooth muscle contraction, etc.
