BIOCHEM MIDTERM

1ST
Week-9 CHEM113

ENZYMES – E. g., In the fermentation process sugar to

• Enzymes are catalysts and are not consumed in the be converted to CO2, therefore in this
reactions reaction sugar is the substrate
• Enzymes are proteins that act as a catalyst for
biochemical reactions THREE IMPORTANT ASPECTS OF THE NAMING PROCESS
• The human body has 1000s of enzymes
• Enzymes are the most effective catalysts known 1. Suffix -ase identifies it as an enzyme
• Most enzymes are globular proteins – E.g., urease, sucrase, and lipase are all
• A few enzymes are now known to be ribonucleic acids enzyme designations
(RNA) – Exception: The suffix -in is still found in the
• Enzymes undergo all the reactions of proteins names of some digestive enzymes, E.g.,
including denaturation trypsin, chymotrypsin, and pepsin
• Enzyme activity is dramatically affected by: 2. Type of reaction catalyzed by an enzyme is often used
– Alterations in pH as a prefix
– E.g., Oxidase - catalyzes an oxidation
– Temperature
reaction,
– Other protein denaturants
– E.g., Hydrolase - catalyzes a hydrolysis
ENZYME STRUCTURE reaction
3. Identity of substrate is often used in addition to the
SIMPLE AND CONJUGATED ENZYMES type of reaction
• Enzymes are of two types: simple enzymes and – E.g. Glucose oxidase, pyruvate carboxylase,
conjugated enzymes and succinate dehydrogenase
• Simple enzyme: composed only of protein (amino acid
chains) SIX MAJOR CLASSES
• Conjugated enzyme: Has a nonprotein part in addition Enzymes are grouped into six major classes based on
to a protein part. the types of reactions they catalyze.
CLASS REACTION CATALYZED
– Apoenzyme: Protein part of a conjugated
Oxidoreductases Reaction catalyzed
enzyme.
Transferases Functional group transfer
– A cofactor : Nonprotein part of a reactions
conjugated enzyme. Hydrolases Hydrolysis reactions
– A holoenzyme is the biochemically active Lyases Reactions involving
conjugated enzyme addition or removal of
– Apoenzyme + cofactor = holoenzyme groups from double
(conjugated enzyme) bonds
Isomerase Isomerization reactions
COFACTORS Ligases Reaction involving bond
• Cofactors provide additional chemically reactive formation coupled with
functional group besides those present in amino acid ATP hydrolysis
side chains of apoenzymes.
• Many cofactors are permanently/temporarily bonded OXIDOREDUCTASE
via covalent bond to the amino acid portion of an
enzyme. • An oxidoreductase enzyme catalyzes an oxidation–
reduction reaction:
• Categories of cofactors
a) Simple metal: Zn2+, Mg2+, Mn2+, Cu and Fe2+ – Oxidation and reduction reactions are
b) Coenzymes : Small organic molecules in a always linked to one another
conjugated enzyme. – An oxidoreductase requires a coenzyme
– FAD (Flavin Adenine Dinucleotide) that is either oxidized or reduced as the
substrate in the reaction.
– NAD (Nicotinamide Adenine
Dinucleotide – E.g., Lactate dehydrogenase is an
NOTE: All the coenzymes are cofactors but all cofactors oxidoreductase that removes hydrogen
cannot be coenzymes. atoms from a molecule.
NOMENCLATURE AND CLASSIFICATION OF ENZYMES
TRANFERASE
• Nomenclature: Are named with reference to their
function
• A transferase is an enzyme that catalyzes the transfer
– Type of reaction catalyzed of a functional group from one molecule to another
– Identity of the substrate • Two major subtypes:
• A substrate is the reactant in an enzyme-catalyzed – Transaminases - catalyze transfer of an
reaction: amino group to a substrate
– The substrate is the substance upon which
the enzyme “acts.”
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Week-9 CHEM113

– Kinases - catalyze transfer of a phosphate

group from adenosine triphosphate (ATP)
to a substrate.

HYDROLASES

• 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:
– Carbohydrase’s 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 bond
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 ACTION
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
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
• 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)
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)
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– 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 SUBSTRATE CONCENTRATION
• Substrate Concentration: At a constant enzyme
FACTORS THAT AFFECT ENZYME ACTIVITY concentration, the enzyme activity increases with
increased substrate concentration.
TEMPERATURE • Substrate saturation: the concentration at which it
• Higher temperature results in higher kinetic energy reaches its maximum rate and all of the active sites
which causes an increase in number of reactant are full
collisions, therefore there is higher activity. • Turnover Number: Number of substrate molecules
• Optimum temperature: Temperature at which the rate converted to product per second per enzyme
of enzyme catalyzed reaction is maximum molecule under conditions of optimum temperature
• Optimum temperature for human enzymes is 37ºC and pH
(body temperature)
• Increased temperature (high fever) leads to decreased
enzyme activity

ENZYME CONCENTRATION
• Enzyme Concentration:
• Enzymes are not consumed in the reactions they
catalyze
• At a constant substrate concentration, enzyme activity
pH increases with increase in enzyme concentration
• pH changes affect enzyme activity – The greater the enzyme concentration, the
• Drastic changes in pH can result in denaturation of greater the reaction rate.
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
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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.
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+.
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 produce large amounts
of an enzyme and plentiful amounts of products if the
processes are not regulated.
• General mechanisms involved in regulation:
– Proteolytic enzymes and zymogens
covalent modification of enzymes
– Feedback control Regulation of enzyme
activity by various substances produced
within a cell
– The enzymes regulated are allosteric
enzymes
MIDTERM
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CHEM113
PROPERTIES OF ALLOSTERIC ENZYMES
• All allosteric enzymes have quaternary structure:
– Composed of two or more protein subunits
• Have at least two 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 form
(zymogens)
– Zymogens are “turned on” at the
appropriate time and place. It is the
inactive precursor of a proteolytic enzyme.
– 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
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COVALENT MODIFICATION OF ENZYMES PENICILLIN

• 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.
PRESCRIPTION DRUGS THAT INHIBIT ENZYME ACTIVITY
• ACE ( angiotensin-converting enzymes) inhibitors
– an octapeptide hormone involved in blood
pressure regulation. Increases blood
pressure by narrowing blood vessel.
– present as zymogen in the form of
angiotensinogen.
example: Lisinopril
ANTIBIOTICS THAT INHIBIT ENZYME ACTIVITY
• An antibiotics 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 penicillin
SULFA DRUGS
• 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
• Accidently discovered by Alexander Fleming in 1928
• Several naturally occurring penicillin 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
Enzyme Conditions Indicated by
Abnormal level
Lactate Heart disease, liver disease
dehydrogenase (LDH)
Creatinine Heart disease
phosphokinase (CPK)
Aspartate Heart disease, liver disease,
aminotransferase (AST) muscle damage
Alanine Heart disease, liver disease,
aminotransferase (ALT) muscle damage
Gamma-glutamyl Heart disease, liver disease
transpeptidase (GGPT)
Alkaline phosphatase Bone disease, liver disease