Lectures 9 and 10 – Enzymes

I. Introduction:

 All reactions in the body are mediated by enzymes. Enzymes act on reactants
called substrates.
 Enzymes are protein catalysts that increase the velocity of a chemical reaction,
and are not consumed during the reaction they catalyze (i.e. no changes in the
overall process).

II. Nomenclature:

 There are 2 systems to name the enzymes:

1) Recommended name: Most commonly used enzyme names

(a) have the suffix "-ase" attached to the substrate of the reaction (for example,
glucosidase urease, sucrase),
(b) The name represents a description of the action performed (for example,
lactate dehydrogenase and adenylyl cyclase).
(c) Some enzymes retain their original trivial names, which give no hint of the
associated enzymic reaction, for example, trypsin and pepsin.

2) Systematic name:

 The International Union of Biochemistry and Molecular Biology (IUBMB)

developed a system of nomenclature in which enzymes are divided into six
major classes.

III. Major Classes of Enzymes:

1) Oxidoreductases:

 In oxidation and reduction reactions, one substrate is oxidized and one is

reduced.

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  In this reaction, lactate is converted to pyruvate, and the cofactor, NAD, is
converted to NADH. In other words, lactate is oxidized, and NAD is reduced.

2) Transferases:

 Transfer of phosphate group to glucose

Hexokinase

3) Hydrolases:

 As shown in the following reactions, these enzymes break single bonds by adding
the elements of water.

4) Lyases:

 They catalyze the cleavage of C-C, C-S, C-N bonds:

Pyruvate Decarboxylase

5) Isomerases:

 The substrate and product of the reaction are isomers. The isomerases (for
example, triose phosphate isomerase, shown below), carry out these
rearrangements.

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 6) Ligases:

 They catalyze formation of bonds between carbon and O, S, N coupled to

hydrolysis of high energy phosphates.
 Synthetases are a subclass of ligases that use the hydrolysis of ATP to drive this
formation. For example

Pyruvate Carboxylase

IV. Properties of Enzymes:

A) Active Site:
 Enzyme molecules contain a special pocket or cleft called the active site.
 The active site contains amino acid side chains that create a three-dimensional
surface complementary to the substrate.
 The active site binds the substrate, forming an enzyme-substrate (ES) complex.
ES is converted to enzyme-product (EP), which subsequently dissociates to
enzyme and product.

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B) Catalytic Efficiency:
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 Most enzyme-catalyzed reactions are highly efficient, proceeding from 10 to
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10 times faster than uncatalyzed reactions.
 Typically, each enzyme molecule is capable of transforming 100 to 1000
substrate molecules into products each second.
 The number of molecules of substrate converted to product per enzyme
molecule per second is called the turnover number.
 For the enzyme urease, the turnover number is 30,000/sec. For the enzyme
carbonic anhydrase, which converts carbon dioxide and water into carbonic
acid, the turnover number is 1 million/sec.

C) Catalytic Specificity:
 Enzymes are highly specific, interacting with one or a few substrates and
catalyzing only one type of chemical reaction. For examples:

 Urease enzyme: urease is a bacterial enzyme converts urea into water and
ammonia. Therefore, urease has specificity in that it can be absolute for one
molecule.

D) Cofactors and Coenzymes:

 Some enzymes associate with a nonprotein cofactor that is needed for
enzymatic activity. Commonly encountered cofactors include metal ions such
as Zn2+ or Fe2+ and organic molecules, known as coenzymes, that are often
derivatives of vitamins. For example, the coenzyme NAD+ contains niacin,
FAD contains riboflavin, and coenzyme A contains pantothenic acid.
 Coenzymes, on the other hand, are usually derived from the vitamins within
the diet, and much of the early biochemistry of nutrition was devoted to
identifying vitamins as well as the diseases associated with vitamin
deficiencies.
 Holoenzyme refers to the enzyme with its cofactor. Apoenzyme refers to the
protein portion of the holoenzyme.
 In the absence of the appropriate cofactor, the apoenzyme typically does not
show biologic activity.
 A prosthetic group is a tightly bound coenzyme that does not dissociate from
the enzyme.

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Biotin:
 Biotin is a coenzyme in carboxylation reactions, in which it serves as a carrier of
carbon dioxide. Biotin is covalently bound to the e-amino groups of lysine
residues of biotin-dependent enzymes.
 Biotin deficiency does not occur naturally because the vitamin is widely
distributed in food.
 Also, a large percentage of the biotin requirement in humans is supplied by
intestinal bacteria.

Figure 1: A) Structure of biotin. B) Biotin

covalently bound to a lysyl residue of a biotin-
dependent enzyme.

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V. Location within the Cell:
 Many enzymes are localized in specific organelles within the cell.
 Such compartmentalization serves to:
a) Isolate the reaction substrate or product from other competing reactions.
b) Provide a favorable environment for the reaction.
c) Organize the thousands of enzymes present in the cell into purposeful pathways.

Figure 2: The intracellular location of some

important biochemical pathways.

VI: Factors Affecting Reaction Velocity:

A. Substrate concentration:
 The rate or velocity of a reaction (v) is the number of substrate molecules
converted to product per unit time; velocity is usually expressed as μmol of
product formed per minute.
 The rate of an enzyme-catalyzed reaction increases with substrate
concentration until a maximal velocity (Vmax) is reached.

B. Temperature:
1. Increase of velocity with temperature: The reaction velocity increases with
temperature until a peak velocity is reached.
2. Decrease of velocity with higher temperature: Further elevation of the
temperature results in a decrease in reaction velocity as a result of temperature-
induced denaturation of the enzyme.

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C. pH
1. Effect of pH on the ionization of the active site:
 The concentration of H+ affects reaction velocity in several ways. First, the
catalytic process usually requires that the enzyme and substrate have specific
chemical groups in either an ionized or unionized state in order to interact.
 For example, catalytic activity may require that an amino group of the enzyme
be in the protonated form (-NH3+). At alkaline pH this group is deprotonated,
and the rate of the reaction, therefore, declines.

2. The pH optimum varies for different enzymes:

 The pH at which maximal enzyme activity is achieved is different for different
enzymes, and often reflects the [H+] at which the enzyme functions in the body.
 For example, pepsin, a digestive enzyme in the stomach, is maximally active at
pH 2, whereas other enzymes, designed to work at neutral pH, are denatured by
such an acidic environment.

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VII: Regulation of Enzyme Activity:

A. Allosteric binding sites:

 Allosteric enzymes are regulated by molecules called effectors that bind
noncovalently at a site other than the active site.
 Effectors that inhibit enzyme activity are termed negative effectors, whereas
those that increase enzyme activity are called positive effectors.

1) Hemotropic effectors: When the substrate itself serves as an effector, the

effect is said to be homotropic. The presence of a substrate molecule at
one site on the enzyme enhances the catalytic properties of the other
substrate.
2) Heterotropic effectors: The effector may be different from the substrate,
in which case the effect is said to be heterotropic.

B. Regulation of enzymes by covalent modification

 Many enzymes may be regulated by covalent modification, most frequently by the
addition or removal of phosphate groups from specific serine, threonine, or
tyrosine residues of the enzyme by a family of enzymes called protein kinases
that use adenosine triphosphate (ATP) as a phosphate donor.
 Phosphorylated form of enzyme may be either active or inactive depending on the
type of enzyme.
 Phosphate groups are cleaved from phosphorylated enzymes by the action of
phosphoprotein phosphatases.

C. Induction and repression of enzyme synthesis:

 Cells can regulate the amount of enzyme present—usually by altering the rate of
enzyme synthesis.
 For example, elevated levels of insulin as a result of high blood glucose levels
cause an increase in the synthesis of key enzymes involved in glucose
metabolism.

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VIII: Inhibition of Enzyme Activity:
 Inhibitor definition: it is any substance that can diminish the velocity of an
enzyme-catalyzed reaction.
1. Irreversible inhibitors bind to enzymes through covalent bonds.
2. Reversible inhibitors typically bind to enzymes through noncovalent bonds,
thus dilution of the enzyme–inhibitor complex results in dissociation of the
reversibly bound inhibitor, and recovery of enzyme activity.

A. Competitive inhibition:
 This type of inhibition occurs when the
inhibitor binds reversibly to the same site that
the substrate would normally occupy and,
therefore, competes with the substrate for that
site.

B. Noncompetitive inhibition
 Noncompetitive inhibition occurs when the
inhibitor and substrate bind at different sites
on the enzyme.
 The noncompetitive inhibitor can bind either
free enzyme or the enzyme-substrate (ES)
complex, thereby preventing the reaction
from occurring.

C: Uncompetitive inhibition:
 It is seen when an inhibitor can bind to the ES complex but not to free E.

D. Enzyme inhibitors as drugs:

 At least half of the ten most commonly dispensed drugs in the United States act as
enzyme inhibitors. For example, the widely prescribed β-lactam antibiotics, such as
penicillin and amoxicillin act by inhibiting enzymes of bacterial cell wall synthesis.
 Drugs may also act by inhibiting extracellular reactions. This is illustrated by
angiotensin-converting enzyme (ACE) inhibitors. They lower blood pressure by
blocking the enzyme that cleaves angiotensin I to form the potent vasoconstrictor,
angiotensin II. These drugs, which include captopril, enalapril, and lisinopril, cause
vasodilation and a resultant reduction in blood pressure.