ENZYMES

BIOCHEMISTRY

BCH 211

ODUGBEMI A. I.
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Enzymes

 Thousands of chemical reactions are proceeding very rapidly at any given instant within all living
cells. Virtually all of these transformations are mediated by enzymes—proteins (and occasionally
RNA) specialized to catalyze metabolic reactions.

 Enzymes form metabolic pathways by which nutrient molecules are degraded, energy is released and
converted into metabolically useful forms, and precursors are generated and transformed to create the
literally thousands of distinctive biomolecules found in any living cell.

 Enzymes are remarkably versatile biochemical catalysts that have in common three distinctive
features: catalytic power, specificity, and regulation.

 Enzymes display enormous catalytic power, accelerating reaction rates as much as 1021 over
uncatalyzed levels, which is far greater than any synthetic catalysts can achieve, and enzymes
accomplish these astounding feats in dilute aqueous solutions under mild conditions of temperature
and pH

 A given enzyme is very selective, both in the substances with which it interacts and in the reaction
that it catalyzes. The substances upon which an enzyme acts are traditionally called substrates.

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Basic Characteristics of Enzymes

 Enzymes do not make anything happen that couldn’t happen on its own, just
makes it happen faster.

 Enzymes lower activation energy

 Enzymes are not used up in reactions. They can be used over and over again

 Enzymes are only needed in small amounts.

 Each enzyme is highly selective about its substrate.

 Enzymes chemically recognize, bind and modify substrates.

 Enzymes are water soluble

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Enzymes Lower Activation Energy

 The difference between the energies of the

reactants (the initial state) and the energies
of the products (the final state) of a reaction
gives the energy change for that reaction,
expressed as the standard free energy
change, or ΔG°.

 Enzymes, like all catalysts, speed up

reactions, but they cannot alter the
equilibrium constant or the free energy
change.

 The reaction rate depends on the free

energy of activation or activation energy
(ΔG°*), the energy input required to initiate
the reaction.

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Enzyme Active Site

The active site of an enzyme is the region that binds the substrates (and the
cofactor, if any).
It also contains the residues that directly participate in the making and
breaking of bonds. These residues are called the catalytic groups.
In essence, the interaction of the enzyme and substrate at the active site
promotes the formation of the transition state.

The active site is a three-dimensional cleft, or crevice, formed by groups

that come from different parts of the amino acid sequence.
The active site takes up a small part of the total volume of an enzyme
Substrates are bound to enzymes by multiple weak attractions –
electrostatic interactions, hydrogen bonds, and van der Waals forces
The specificity of binding depends on the precisely defined arrangement of
atoms in an active site.

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Model of Catalysis

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Enzyme Naming

 Since earlier days to still date, fanciful names such as pepsin, chymotrypsin, etc
were used to name enzymes.

 Later the suffix “ase” to the substrate was used to name enzymes. For example the
enzymes lactase acts upon the lactate and produces glucose and galactose.

 The above method is known as “trivial naming” of enzymes.

 Now the two name system is used. The substrate or product of enzymes and the
reaction involved. For example, Glucose-6-phosphate dehydrogenase

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Enzyme Classification

 Currently enzymes are grouped into six functional classes by the International Union of
Biochemists and Molecular Biology (IUBMB).

 As per the IUBMB system, each enzyme name starts with EC (enzyme class) followed by
4 digits.

 The first digit represents the class, the second digit strands for the subclass, the third digit
represents the sub-subclass or subgroup and the fourth digit provides the particular
enzyme.

 For example, the enzyme commonly called “hexokinase” is designated “ATP:D-hexose-6-

phosphotransferase E.C. 2.7.1.1.”

 This identifies hexokinase as a member of class 2 (transferases), subclass 7 (transfer of a

phosphoryl group), sub-subclass 1 (alcohol is the phosphoryl acceptor).

 Finally, the term “hexose-6” indicates that the alcohol phosphorylated is that of carbon six
of a hexose.
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Enzyme Classification
1. Oxidoreductases catalyze oxidations and reductions e.g. Lactate dehydrogenase

2. Transferases catalyze transfer of functional groups such as methyl, glycosyl, amino or phosphate
groups from a donor molecule to an acceptor molecule e.g. Aminotransferase. Kinases are specialized
transferases that regulate metabolism by transferring phosphate from ATP to other molecules.

3. Hydrolases catalyze the hydrolytic cleavage of C – C, C – O, C – N, P – O, and certain other bonds,

including acid anhydride bonds. Basically adding water across a bond, hydrolyzing it e.g. Acetyl
choline esterase.

4. Lyases catalyze cleavage of C – C, C – O, C – N, and other bonds by elimination, leaving double

bonds, and also add groups to double bonds. Add water, ammonia or carbon dioxide across double
bonds, or remove these elements to produce double bonds. e.g. Aldolase.

5. Isomerases catalyze geometric or structural changes within a single molecule. They carry out many
kinds of isomerization: L to D isomerizations, mutase reactions and others e.g. Triose phosphate
isomerase.

6. Ligases catalyze the joining together of two molecules with the use of energy from ATP. e.g. Acetyl
CoA carboxylase
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Enzyme Classification

 Enzymes are also classified on the basis of their composition.

 Enzymes composed wholly of protein are known as simple enzymes in contrast to complex
enzymes, which are composed of protein plus a relatively small organic molecule. Complex
enzymes are also known as holo-enzymes. The protein component is termed apoenzyme.

 The nonprotein component of an enzyme may be as simple as a metal ion or as complex as a small
non-protein organic molecule.

 Enzymes that require a metal in their composition are known as metalloenzymes.

 Based on requirement of ATP, enzymes are further classified into two types namely synthetases and
synthase. Synthetases are ATP-dependent enzymes catalyzing biosynthetic reactions. Synthetases
are enzyme belong to the class 6 (Ligases) e.g. Carbamoyl phosphate synthetase. The enzyme class
other than ligases includes synthases. Synthases group of enzymes involves in catalyzing
biosynthetic reactions that do not require ATP directly. Enzymes such as glycogen synthase and
Alanine synthase are examples of synthase group.

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Coenzymes

 Enzymes may be simple proteins, or complex enzymes. A complex enzyme contains a non-
protein part, called co-enzymes.

 The combined form of protein and the co-enzyme are called as holo-enzyme.

 Coenzymes are basically the organic cofactors required for enzyme activities.

 Co-enzymes are further divided into two groups. The first group of co-enzymes are also
called as co-substrates. The second group of co-enzymes are called prosthetic group.

 The functional role of coenzymes is to act as transporters of chemical groups from one
reactant to another.

 Many physiological coenzymes are derivatives of vitamins obtained from diet.

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Factors Affecting Enzyme Activity

Substrate concentration

 Reaction velocity of an enzymatic process increases with constant enzyme concentration and increase
in substrate concentration.

 The velocity (V) is expressed in micromoles of substrate converted per minute.

 As the concentration of substrate increases, the velocity of the reaction increases. Continued increase in
substrate concentration may lead to a reduction in rate of the reaction and leads to flattened curve. The
maximum velocity obtained from a enzymatic reaction is called as Vmax.

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Factors Affecting Enzyme Activity
Enzyme concentration
As there is optimal substrate concentration, rate of an enzymatic reaction or velocity (V) is directly
proportional to the enzyme concentration.

Product concentration
The rate of reaction is slowed, stopped or even reversed with increase in product concentration.
This phenomena can be better explained by the equation

In the above equation, in case of absence of the enzyme E3, the product C will accumulate. Enzymatic
activity of E2 will be inhibited with accumulation of the product C. In such inborn error of one enzyme
will block the whole pathway.
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Temperature
 Velocity of enzymatic reaction increases with temperature of
the medium which they are most efficient and the same is
termed as optimum temperature.
 As temperatures increases it leads to denaturation; a molecular
arrangement which causes a loss of the active sites of the
enzyme surfaces and results in a loss of efficiency.

Hydrogen ion concentration (pH)

 Like temperature, all enzymes have a optimum pH, at which the
enzymatic activity will be at maximum.
 Many enzymes are most efficient in the region of pH 6-7, which
is the pH of the cell. Outside this range, enzyme activity drops
off very rapidly.
 Reduction in efficiency caused by changes in the pH is due to
changes in the degree of ionization of the substrate and enzyme.
 Highly acidic or alkaline conditions bring about a denaturation
and subsequent loss of enzymatic activity.
 Some exceptions such as pepsin (with optimum pH 1-2),
alkaline phosphatase (with optimum pH 9-10) and acid
phosphatase (with optimum pH 4-5).
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Presence of activators
Presence of certain inorganic ions increases the activity of enzymes.
The best examples are chloride ions activated salivary amylase and calcium activated lipases.

Presence of inhibitor
The catalytic enzymatic reaction may be inhibited by substances which prevent the formation
of a normal enzyme-substrate complex.
Enzyme inhibitor plays a vital role in clinical utility

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Distinction between reversible inhibitors. (A) Enzyme–substrate complex; (B) a competitive inhibitor
binds at the active site and thus prevents the substrate from binding; (C) an uncompetitive inhibitor
binds only to the enzyme–substrate complex; (D) a noncompetitive inhibitor does not prevent the
substrate from binding.
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Regulation of Enzyme Activity
The activity of enzymes must often be regulated so that they function at the proper time and place.

 Allosteric Control. Allosteric enzymes contain distinct regulatory sites and multiple functional sites. The
binding of small signal molecules at regulatory sites is a significant means of controlling the activity of
these proteins. Moreover, allosteric proteins show the property of cooperativity: activity at one
functional site affects the activity at others.
 Multiple Forms of Enzymes. Isozymes, or isoenzymes, provide an avenue for varying regulation of the
same reaction at distinct locations or times to meet the specific physiological needs in the particular tissue
at a particular time.
 Reversible Covalent Modification. The catalytic properties of many enzymes are markedly altered by
the covalent attachment of a modifying group, most commonly a phosphoryl group. ATP serves as the
phosphoryl donor in these reactions, which are catalyzed by protein kinases. The removal of phosphoryl
groups by hydrolysis is catalyzed by protein phosphatases.
 Proteolytic Activation. This irreversibly convert an inactive enzyme into an active one. Many enzymes
are activated by the hydrolysis of a few peptide bonds or even one such bond in inactive precursors called
zymogens or proenzymes. This regulatory mechanism generates digestive enzymes such as
chymotrypsin, trypsin, and pepsin. Blood clotting is due to a remarkable cascade of zymogen activations.
 Controlling the Amount of Enzyme Present. Enzyme activity can also be regulated by adjusting the
amount of enzyme present. This important form of regulation usually takes place at the level of
transcription.
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Michaelis-Menten Equation

 The Michaelis-Menten equation illustrates in mathematical terms the relationship

between initial reaction velocity Vi and substrate concentration [S], shown graphically.

 The Michaelis constant Km is the substrate concentration at which Vi is half the maximal
velocity (Vmax/2) attainable at a particular concentration of enzyme.
 Km thus has the dimensions of substrate concentration.

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