BACHELOR OF SCIENCE IN MEDICAL

LABORATORY SCIENCE:

BIOCHEMISTRY LECTURE
COURSE MODULE COURSE UNIT SESSION WEEK
1 3 2 1
Enzymes

 Read course and unit objectives

 Read study guide prior to online attendance
 Read required learning resources; refer to unit
terminologies for jargons
 Proactively participate in online discussions
 Participate in weekly discussion board (Canvas)
 Answer and submit course unit tasks

At the end of this unit, the students are expected to:

Cognitive:
1. List accordingly the different classification of enzymes
2. Describe correctly different enzymes structures
3. Enumerate accurately the factors that affect enzyme activity
4. Describe fittingly the models of enzymes activities
5. Identify appropriately the enzymes which inhibits enzyme activity
Affective:
1. Inculcate correctly importance of enzymes in different biochemical reactions in the body
2. Listen attentively during online discussions
3. Demonstrate tact and respect when challenging other people’s opinions and ideas
4. Accept comments and reactions of classmates on one’s opinions openly and graciously.

Psychomotor:
1. Participate actively during online discussions
2. Confidently express personal opinion and thoughts in online discussions

Stoker, S (2015). General, Organic, & Biological Chemistry 7 th Edition, Enzymes (pp. 754-776).
New York: Cengage Learning (30 item Assessment Task will be coming from this reference).

ENZYMES

General Characteristics of Enzymes

Enzymes are catalysts and are not consumed in the reactions
Enzymes are proteins that act as a catalyst for biochemical reactions
The human body has 1000s of enzymes
Enzymes are the most effective catalysts known
Most enzymes are globular proteins
A few enzymes are now known to be ribonucleic acids (RNA)
Enzymes undergo all the reactions of proteins including denaturation

Enzyme activity is dramatically affected by:

 Alterations in pH
 Temperature
 Other protein denaturants
Enzyme Structures
Simple and Conjugated Enzymes

Enzymes are of two types: simple enzymes and conjugated enzymes

Simple enzyme: composed only of protein (amino acid chains)
Conjugated enzyme: Has a nonprotein part in addition to a protein part.
 Apoenzyme: Protein part of a conjugated enzyme.
 A cofactor : Nonprotein part of a conjugated enzyme.
 A holoenzyme is the biochemically active conjugated enzyme
 Apoenzyme + cofactor = holoenzyme (conjugated enzyme)

Apoenzyme
Apoenzyme is the protein part of the enzyme which lacks the cofactor.
Apoenzyme is catalytically inactive and incomplete and determines the specificity of this system for
a substrate.
Cofactors bind tightly or loosely with the apoenzyme to convert apoenzyme into holoenzyme.
Once the cofactor is removed from the holoenzyme, it is converted again into apoenzyme, which is
inactive and incomplete.

[Figure 1. This image shows how a holoenzyme is formed with the binding of cofactor to a apoenzyme. Image downloaded from
www.differencebetween.com/difference-between-holoenzyme-and-vs-apoenzyme/#Apoenzyme]

Cofactors
Cofactors are important for the chemically reactive enzymes
Cofactors are small organic molecules or Inorganic ions
 Organic molecule cofactors: also called as co-enzymes or co-substrates
 Co-enzymes/co-substrates are derived from dietary vitamins
 Inorganic ion cofactors
 Typical metal ion cofactors - Zn2+, Mg2+, Mn2+, and Fe2+
 Nonmetallic ion cofactor - Cl-
  Inorganic ion cofactors derived from dietary minerals

Nomenclature and Classification of Enzymes

Nomenclature: Most commonly named with reference to their function
 Type of reaction catalyzed
 Identity of the substrate
A substrate is the reactant in an enzyme-catalyzed reaction:
 The substrate is the substance upon which the enzyme “acts.”
 E. g., In the fermentation process sugar to be converted to CO2, therefore in this reaction
sugar is the substrate

Three Important Aspects of the Naming Process

Suffix -ase identifies it as an enzyme
 E.g., urease, sucrase, and lipase are all enzyme designations
 Exception: The suffix -in is still found in the names of some digestive enzymes, E.g.,
trypsin, chymotrypsin, and pepsin
Type of reaction catalyzed by an enzyme is often used as a prefix
 E.g., Oxidase - catalyzes an oxidation reaction,
 E.g., Hydrolase - catalyzes a hydrolysis reaction
Identity of substrate is often used in addition to the type of reaction
 E.g. Glucose oxidase, pyruvate carboxylase, and succinate dehydrogenase

Six Major Classes

Enzymes are grouped into six major classes based on the types of reactions they catalyze

Class Reaction Catalyzed

1. Oxidoreductases Oxidation-reductions

2. Transferases Functional group transfer reactions
 3. Hydrolases Hydrolysis reactions

Reactions involving addition or removal of groups form double bo
4. Lyases
nds

5. Isomerase Isomerisation reactions

6. Ligases Reactions involving bond formation coupled with ATP hydrolysis

Oxidoreductase
An oxidoreductase enzyme catalyzes an oxidation–reduction reaction:
 Oxidation and reduction reactions are always linked to one another
 An oxidoreductase requires a coenzyme that is either oxidized or reduced as the substrate
in the reaction.
 E.g., Lactate dehydrogenase is an oxidoreductase and the reaction catalyzed is shown
below

Transferase
A transferase is an enzyme that catalyzes the transfer of a functional group from one molecule to
another
 Two major subtypes:
 Transaminases - catalyze transfer of an amino group to a substrate
 Kinases - catalyze transfer of a phosphate group from adenosine triphosphate (ATP) to a
substrate

Hydrolase
A hydrolase is an enzyme that catalyzes a hydrolysis reaction
The reaction involves addition of a water molecule to a bond to cause bond breakage
Hydrolysis reactions are central to the process of digestion:
  Carbohydrases hydrolyze glycosidic bonds in oligo- and polysaccharides (see reaction
below)
 Proteases effect the breaking of peptide linkages in proteins,
 Lipases effect the breaking of ester linkages in triacylglycerols

Lyase
A lyase is an enzyme that catalyzes the addition of a group to a double bond or the removal of a
group to form a double bond in a manner that does not involve hydrolysis or oxidation
 Dehydratase: effects the removal of the components of water from a double bond
 Hydratase: effects the addition of the components of water to a double bonds

Isomerase and Ligase

An isomerase is an enzyme that catalyzes the isomerization (rearrangement of atoms) reactions.
A ligase is an enzyme that catalyzes the formation of a bond between two molecules involving ATP
hydrolysis:
 ATP hydrolysis is required because such reactions are energetically unfavorable
 Require the simultaneous input of energy obtained by a hydrolysis of ATP to ADP

Models of Enzyme Actions

Enzyme Active Site
The active site: Relatively small part of an enzyme’s structure that is actually involved in catalysis:
 Place where substrate binds to enzyme
 Formed due to folding and bending of the protein.
 Usually a “crevice like” location in the enzyme
 Some enzymes have more than one active site
[Figure 2. This image show parts of the enzyme. Image from Stoker, S (2015).General, Organic, & Biological Chemistry 7th Edition, Enzymes .
New York: Cengage Learning, page 781]

Enzyme Substrate Complex

Needed for the activity of enzyme
Intermediate reaction species formed when substrate binds with the active site
Orientation and proximity is favorable and reaction is fast

Two Models for Substrate Binding to Enzyme

Lock-and-Key model:
 Enzyme has a pre-determined shape for the active site
 Only substrate of specific shape can bind with active site

[Figure 3. This image shows enzyme binding through Lock-and-key model of the enzyme. Image from Stoker, S (2015).General, Organic, &
Biological Chemistry 7th Edition, Enzymes . New York: Cengage Learning, page 781]

Induced Fit Model:

 Substrate contact with enzyme will change the shape of the active site
 Allows small change in space to accommodate substrate (e.g., how a hand fits into a
glove)
[Figure 4. This image shows enzyme binding through Induced fit model of the enzyme. Image from Stoker, S (2015).General, Organic, &
Biological Chemistry 7th Edition, Enzymes . New York: Cengage Learning, page 781]

Forces That Determine Substrate Binding

H-bonding
Hydrophobic interactions
Electrostatic interactions

Enzyme Specificity
Absolute Specificity:
 An enzyme will catalyze a particular reaction for only one substrate
 This is most restrictive of all specificities (not common)
 E.g., Urease is an enzyme with absolute specificity
Stereochemical Specificity:
 An enzyme can distinguish between stereoisomers.
 Chirality is inherent in an active site (amino acids are chiral compounds)
 L-Amino-acid oxidase - catalyzes reactions of L-amino acids but not of D-amino acids.
Group Specificity:
 Involves structurally similar compounds that have the same functional groups.
 E.g., Carboxypeptidase: Cleaves amino acids one at a time from the carboxyl end of the
peptide chain
Linkage Specificity:
 Involves a particular type of bond irrespective of the structural features in the vicinity of the
bond
 Considered most general of enzyme specificities
 E.g., Phosphatases: Hydrolyze phosphate–ester bonds in all types of phosphate esters

Factors that Affect Enzymes Activity

Temperature
Higher temperature results in higher kinetic energy which causes an increase in number of
reactant collisions, therefore there is higher activity.
Optimum temperature: Temperature at which the rate of enzyme catalyzed reaction is maximum
Optimum temperature for human enzymes is 37ºC (body temperature)
Increased temperature (high fever) leads to decreased enzyme activity

[Figure 5. This image shows effect of temperature to enzymatic reaction. Image from Stoker, S (2015).General, Organic, & Biological
Chemistry 7th Edition, Enzymes . New York: Cengage Learning, page 784]

pH
pH changes affect enzyme activity
Drastic changes in pH can result in denaturation of proteins
Optimum pH: pH at which enzyme has maximum activity
Most enzymes have optimal activity in the pH range of 7.0 - 7.5
 Exception: Digestive enzymes
 Pepsin: Optimum pH = 2.0
 Trypsin: Optimum pH = 8.0
[Figure 6. This image shows effect of pH to enzymatic reaction. Image from Stoker, S (2015).General, Organic, & Biological Chemistry 7 th
Edition, Enzymes . New York: Cengage Learning, page 784]

Substracte Concentration
Substrate Concentration: At a constant enzyme concentration, the enzyme activity increases with
increased substrate concentration.
Substrate saturation: the concentration at which it reaches its maximum rate and all of the active
sites are full
Turnover Number: Number of substrate molecules converted to product per second per enzyme
molecule under conditions of optimum temperature and pH

[Figure 7. This image shows effect of substrate concentration to enzymatic reaction. Image from Stoker, S (2015).General, Organic, &
Biological Chemistry 7th Edition, Enzymes . New York: Cengage Learning, page 784]

Enzyme Concentration
Enzyme Concentration:
Enzymes are not consumed in the reactions they catalyze
At a constant substrate concentration, enzyme activity increases with increase in enzyme
concentration
 The greater the enzyme concentration, the greater the reaction rate.

Enzyme Inhibition
Enzyme Inhibitor: a substance that slows down or stops the normal catalytic function of an enzyme
by binding to it.
Competitive Inhibitors: Compete with the substrate for the same active site
 Will have similar charge & shape
 Noncompetitive Inhibitors: Do not compete with the substrate for the same active site
 Binds to the enzyme at a location other than active site

Reversible Competitive Inhibition

A competitive enzyme inhibitor: resembles an enzyme substrate in shape and charge
Binds reversibly to an enzyme active site and the inhibitor remains unchanged (no reaction occurs)
The enzyme - inhibitor complex formation is via weak interactions (hydrogen bonds, etc.).
Competitive inhibition can be reduced by simply increasing the concentration of the substrate.

[Figure 8. This image shows binding of inhibitor to the active site. Image from Stoker, S (2015).General, Organic, & Biological Chemistry 7 th
Edition, Enzymes . New York: Cengage Learning, page 788]

Reversible Noncompetitive Inhibition

A noncompetitive enzyme inhibitor decreases enzyme activity by binding to a site on an enzyme
other than the active site.
Causes a change in the structure of the enzyme and prevents enzyme activity.
Increasing the concentration of substrate does not completely overcome inhibition.
Examples: Heavy metal ions Pb2+, Ag+, and Hg2+.
[Figure 9. This image shows competitive inhibition. Image from Stoker, S (2015).General, Organic, & Biological Chemistry 7 th Edition,
Enzymes . New York: Cengage Learning, page 788]

Irreversible Inhibition
An irreversible enzyme inhibitor inactivates enzymes by forming a strong covalent bond with the
enzyme’s active site.
 The structure is not similar to enzyme’s normal substrate
 The inhibitor bonds strongly and increasing substrate concentration does not reverse the
inhibition process
 Enzyme is permanently inactivated.
 E.g., Chemical warfare agents (nerve gases) and organophosphate insecticides

Regulation of Enzyme Activity

Cellular processes continually produces large amounts of an enzyme and plentiful amounts of
products if the processes are not regulated.
General mechanisms involved in regulation:
 Proteolytic enzymes and zymogenscovalent modification of enzymes
 Feedback control Regulation of enzyme activity by various substances produced within a
cell
 The enzymes regulated are allosteric enzymes

Properties of Allosteric Enzymes

All allosteric enzymes have quarternary structure:
 Composed of two or more protein chains
Have at least two of binding sites:
 Substrate and regulator binding site
Active and regulatory binding sites are distinct from each other:
 Located independent of each other
Shapes of the sites (electronic geometry) are different
Binding of molecules at the regulatory site causes changes in the overall three dimensional
structure of the enzyme:
 Change in three dimensional structure of the enzyme leads to change in enzyme activity
 Some regulators increase enzyme activity – activators
 Some regulators decrease enzyme activity - inhibitors

Feedback Control
Feedback Control: A process in which activation or inhibition of the first reaction in a reaction
sequence is controlled by a product of the reaction sequence.
Regulators of a particular allosteric enzyme may be:
 Products of entirely different pathways of reaction within the cell
 Compounds produced outside the cell (hormones)

Proteolytic Enzymes and Zymogens

2nd mechanism of regulating enzyme activity:
 Production of enzymes in an inactive forms (zymogens)
 Zymogens are “turned on” at the appropriate time and place
 Example: proteolytic enzymes: Most digestive and blood-clotting enzymes are proteolytic
enzymes
 Hydrolyze peptide bonds in proteins
 Proteolytic enzymes are generated in an inactive form and then converted to their active
form

[Figure 10. This image shows conversion of zymogen to a proteolytic enzyme. Image from Stoker, S (2015).General, Organic, & Biological
Chemistry 7th Edition, Enzymes . New York: Cengage Learning, page 792]
Covalent Modification of Enzymes
3rd Mechanism for regulation of enzyme activity
Covalent modification: A process in which enzyme activity is altered by covalently modifying the
structure of the enzyme:
 Involves adding or removing a group from an enzyme
Most common covalent modification: addition and removal of phosphate group:
 Phosphate group is often derived from an ATP molecule.
 Addition of the phosphate (phosphorylation) catalyzed by a Kinase enzyme
 Removal of the phosphate group (dephosphorylation) catalyzed by a phosphatase
enzyme.
 Phosphate group is added to (or removed from) the R group of a serine, tyrosine, or
threonine amino acid residue in the enzyme regulated.

Antibiotics that Inhibits Enzyme Actions

An anitibiotic is a substance that kills bacteria or inhibits their growth
Antibiotics usually inhibit specific enzymes essential to life processes of bacteria
Two families of antibiotics considered in this discussion are sulfa drugs and penicillins

Sulfa Grugs
Many derivatives of sulfanilamide collectively called sulfa drugs exhibit antibiotic activities
Sulfanilamide is structurally similar to PABA (p-aminobenzoic acid)
Many bacteria need PABA to produce coenzyme, folic acid
Sulfanilamide is a competitive inhibitor of enzymes responsible for converting PABA to folic acid in
bacteria
Folic acid deficiency retards bacterial growth and that eventually kills them
Sulfa drugs don’t affect humans because we absorb folic acid from our diet

Penicillins
Accidently discovered by Alexander Fleming in 1928
Several naturally occurring penicillins and numerous synthetic derivatives have been produced
All have structures containing a four-membered Beta-lactam ring fused with a five-membered
thiazolidine ring
Selectively inhibits transpeptidase by covalent modification of serine residue
Transpeptidase catalyzes the formation of peptide cross links between polysaccharides strands in
bacterial cell walls

Cipro
The antibiotic ciprofloxacin hydrochloride (Cipro for short)
Considered the best broad-spectrum antibiotics because it is effective against skin and bone
infections as well as against infections involving the urinary, gastrointestinal, and respiratory
systems
It is the drug of choice for treatment of traveler’s diarrhea
Bacteria are slow to acquire resistance to Cipro.
Biochemical threats associated with terrorism has thrust Cipro into the spotlight because it is
effective against anthrax.

Medical Uses of Enzymes

Diagnose certain diseases:
 Enzymes produced in certain organ/tissues if found in blood may indicate certain damage
to that organ/tissue

ACTIVE SITE- a region on the surfa e of an enzyme whose shape permits binding only of a
specific molecular substrate that then undergoes catalysis.
ALLOSTERIC SITE- relating to, undergoing, or being a change in the shape and activity of protein
(such as enzyme) that results from combination with another substance at a point other than the
chemically activie site.
CATALYSIS- a modification and especially increase in the rate of a chemical reaction induced by
material unchanged chemically at the end of the reaction.
SPECIFICITY- the condition of participating in or catalyzing only one or a few chemical reactions.
Worthington Biochemical Corporation (2020), Introduction to Enzymes. Retrieved May 13, 2020
from Worthington website:
http://www.worthington-biochem.com/introBiochem/default.html

Study Questions

 How are enzymes differ in their mechanism of reactions with other enzymes?
 How are enzymes measured?
 What are the significance of enzymes in the following:
 Preventive Medicine
 Diagnosis of diseases
 Treatment monitoring
 Food industries

Stoker, S (2015). General, Organic, & Biological Chemistry 7 th

Edition, Enzymes (pp. 607-650). New York: Cengage Learning

Raymond, K (2016). General, Organic, & Biological Chemistry 3 rd

Edition, An Integrated Approach with Survey Flyer, Enzymes (pp
208-225). New York: Hohn Wiley & Sons Incorporated