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Enzymes
Definition of an enzyme
Enzyme is
protein
catalyst (i.e. increase the rate of reactions)
NOT changed during the reaction
Enzymes direct all metabolic reactions occurring in the cells
Classification of enzymes
1- Oxidoreductase:
catalyses oxidation-reduction reactions
2-Transferases:
catalyses transfer of C-, N- or P- containing groups
3-Hydrolases:
catalyses breakdown of bonds by addition of water (hydrolysis)
4- Lyases:
catalyses breakdown of C – C , C-S & C-N bonds
5- Isomerase:
catalyses racemization of isomers (D- to L- or L- to D-)
6- Ligases:
catalyses formation of bonds between carbon and O, S, N
Mechanism of action of enzymes:
lock & key model
An enzyme binds a substrate in a region called the active site
Enzyme active site is rigid (stable, not malleable)
Only certain substrate can fit the active site of the enzyme
(i.e. the enzyme can act on only one or few substrates. i.e. specific)
P
S
+
S
P
E
+
S
ES complex
E
+
P
Mechanism of enzyme action:
induced fit model
Active site of the enzyme adjust s its shape to bind the substrate.
Enzyme structure is flexible (not rigid)
This mechanism gives increased range of substrate specificity (many
types of substrates can bind the same enzyme)
P
S
SS
P
E
+ S
ES complex
E + P
Properties of enzymes
1- All Enzymes are Proteins (except ribozymes that are RNA)
2- Active Site
Each enzyme molecules contain a special pocket called the active site.
The substrate bind with the active site to form enzyme substrate-complex
(ES) which is converted to enzyme-product complex (EP).
EP dissociates to enzyme and product
3-Catalytic Efficiency:
Each enzyme molecule is capable of transforming 100 – 1000 substrate
molecules into product each second
Properties of enzymes (cont.)
4- Specificity:
Enzymes are highly specific
i.e. each enzyme interacts with one or few type of substrates
and catalyzes one type of reactions
5- Holoenzymes:
Some enzymes require molecules other than proteins for enzymic activity.
Holoenzymes: apoenzyme (protein part) + nonprotein component
Apoenzymes are inactive without the nonprotein component
non-protein part:
1- a metal ion as zinc or ferrous ions (called cofactors)
2- small organic molecules (called coenzymes)
Coenzymes :
may be only transiently associated with the enzyme
or may be permanently associated with the enzyme (prosthetic group)
Coenzymes may be derived from vitamins. (e.g. NAD+ contains niacin)
Properties of enzymes (cont.)
6- Regulation:
Enzyme activity can be regulated by activation or inhibition.
7- Location within the cell:
Many enzymes are localized in specific organelles within the cell.
This serves to isolate the reaction substrate or product from other opposite
reactions.
Factors affecting enzyme activity:
1- Temperature
Optimum temperature
Reaction
Rate
1- Little activity at low temperature
2- Rate of activity increases with temperature
3- Most active at optimum temperatures
(usually 37°C in all enzymes of humans)
4- Activity is lost with denaturation at high
temperatures
Low
High
Temperature
Factors affecting enzyme activity:
2- pH
Maximum activity at optimum pH
where:
R groups of amino acids have proper charge
Tertiary structure of enzyme is intact
Most enzymes lose activity in very low or high pH (due to loss of tertiary structure i.e. denatured)
Each enzyme has its own optimum pH
Optimum pH
Reaction
Rate
pH
3
5
7 9 11
Each enzyme has its own optimum pH
Each enzyme has its own optimum pH
level of blood
Factors affecting enzyme activity:
3- Substrate concentration
Increasing substrate concentration increases the rate of reaction (with
enzyme concentration is constant)
Maximum activity reached when all of enzyme molecules combine with
substrate
Maximum Activity
Reaction
Rate
Substrate Concentration
Factors affecting enzyme activity:
4- Enzyme concentration
Increasing enzyme concentration increases the rate of reaction
(with substrate concentration is constant)
Reaction
Rate
Enzyme Concentration
Factors affecting enzyme activity:
5- Inhibition of enzyme activity
Inhibitor:
is an any substance that can diminish the velocity of a reaction which is catalyzed
by an enzyme.
Reversible Inhibitors: bind to enzymes through non-covalent bonds
Irreversible Inhibitor: bind to enzymes through covalent bonds
Types of inhibitors according to site of binding of inhibitor:
Competitive Inhibitor :
inhibitor binds at the active site of the enzyme
Noncompetitive Inhibitor: inhibitor binds to a site other than active site
Km of enzyme
Km (Michaelis Constant) of an enzyme is numerically equal to the substrate
concentration at which the velocity of reaction is equal to 1/2 Vmax
Km is the substrate concentration at which 1/2 maximal velocity is reached
If Km is small,
the substrate concentration required for the reaction to reach 1/2 maximal
velocity is small.
i.e. the enzyme has a high affinity for the substrate.
If Km is large,
the substrate concentration required for the reaction to reach
1/2 maximal velocity is large.
i.e. the enzyme has a low affinity for the substrate
Competitive inhibitors
A competitive inhibitor:
•
has a structure similar to substrate of the enzyme
•
occupies active site of the enzyme
•
competes with substrate for occupying the active site of the enzyme
•
increases Km of the enzyme to its substrate.
i.e. more substrate is required to reach ½ Vmax
•
effect can be reversed by increasing substrate concentration.
Noncompetitive inhibitors
A noncompetitive inhibitor:
•
Has a structure different from the substrate
•
Binds to the enzyme at a site different from active site
i.e. does not compete with the substrate of the enzyme for the active site of the
enzyme
•
Changes the shape of enzyme and thus active site shape is changed.
Accordingly, substrate cannot fit in the altered active site
So, no reaction occurs
•
Effect is not reversed by adding more substrate
•
Does not change Km of the enzyme (i.e. increase of substrate concentration will not
lead to reaching of ½ Vmax).
Regulation of enzyme activity
The regulation of the activity of enzyme is essential for coordinating the metabolic
processes.
Types of regulation:
1- General: (occurs in all types of enzymes in the body)
increasing substrate concentration will lead to increase activity of the
enzyme
2-Special regulatory mechanisms: (not all enzymes of the body)
i- Allosteric effectors
ii- Covalent modification
iii. Increase or decrease rate of enzyme synthesis
1- Allosteric
effectors
•
•
Allosteric binding sites are sites on the enzyme different from the active site.
Binding of an allosteric effectors to this site will make changes in shape of the
whole enzyme with an effect on activity.
•
So, the activity may be decreased (positive allosteric effector) or decreased
(negative allosteric effector).
2- Covalent
modification
1- Many enzymes may be regulated by addition of phosphate groups to the
enzyme (modification by phosphorylation)
Addition of phosohate group may cause activation or inactivation of the
enzyme
2- Some enzymes are released as an inactive form (zymogen)
By removal of a part of the enzyme (modification), it will be active.
3- Increasing or decreasing rate of enzyme
synthesis
•
By this mechanism cells regulate the amount of enzyme by changing the rate
of enzyme synthesis
•
The increase (induction) or decrease (repression) of enzyme synthesis leads to
change total amount of active site.
•
This mechanism is slow (takes from hours or days)
Medical importance of enzymes of blood
Blood enzymes can be classified into two major groups:
1- Enzymes that have functions in blood (a smaller group):
They are present in blood in high amounts
Example: liver secretes zymogens (inactive precursors) involved in
blood coagulation.
2-Enzymes that do not have functions in blood (a large number)
These enzymes are released from cells during normal cell turn over.
They have functions in cells (intracellular)
BUT: they do not have a function in blood
In healthy individuals, levels of these enzymes are constant .
So, the presence of elevated enzyme level in blood may indicate tissue
damage that leads to increase in release of these enzymes.
Medical importance of enzymes of blood
(cont.)
•
Many diseases that cause tissue damage results in increased release of
intracellular enzymes into plasma (blood)
So, the enzyme levels in blood are measured for diagnosis of these diseases
•
Diseases of the heart, liver, skeletal muscles and other tissues are diagnosed by
an elevation of a blood enzymes.
•
The level of elevation of an enzyme correlates with the extent of tissue
damage in any of these organs.
•
Some enzymes may be available in high amount in only one organ
So, the elevation of blood levels of these enzymes are diagnostic for
diseases of this organ only. (specific)
Example: Alanine aminotransferase (ALT) enzyme elevation in blood
indicates disease of the liver (specific for liver cells).