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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1ST
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
ENZYME SUBSTRATE COMPLEX
LYASE • Needed for the activity of enzyme
• Intermediate reaction species formed when substrate
• A lyase is an enzyme that catalyzes the addition of a binds with the active site
group to a double bond or the removal of a group to • Orientation and proximity is favorable and reaction is
form a double bond in a manner that does not involve fast
hydrolysis or oxidation
– Dehydratase: effects the removal of the TWO MODELS FOR SUBSTRATE BINDING TO ENZYME
components of water from a double bond • Lock-and-Key model:
– Hydratase: effects the addition of the – Enzyme has a pre-determined shape for the
components of water to a double bond active site
– Only substrate of specific shape can bind
ISOMERASE AND LIGASE with active site
• Induced Fit Model:
• An isomerase is an enzyme that catalyzes the – Substrate contact with enzyme will change
isomerization (rearrangement of atoms) reactions. the shape of the active site
• A ligase is an enzyme that catalyzes the formation of – Allows small change in space to
a bond between two molecules involving ATP accommodate substrate (e.g., how a hand
hydrolysis: fits into a glove)
– ATP hydrolysis is required because such
reactions are energetically unfavorable FORCES THAT DETERMINE SUBSTRATE BINDING
– Require the simultaneous input of energy • H-bonding
obtained by a hydrolysis of ATP to ADP • Hydrophobic interactions
• Electrostatic interactions
MODELS OF ENZYME ACTION
ENZYME SPECIFICITY
ENZYME ACTIVE SITE
• Absolute Specificity:
• The active site: Relatively small part of an enzyme’s
structure that is actually involved in catalysis: – An enzyme will catalyze a particular
reaction for only one substrate
– Place where substrate binds to enzyme
– This is most restrictive of all specificities
– Formed due to folding and bending of the
(not common)
protein.
– E.g., Urease is an enzyme with absolute
– Usually a “crevice like” location in the
specificity
enzyme
• Stereochemical Specificity:
– Some enzymes have more than one active
site – An enzyme can distinguish between
stereoisomers.
– Chirality is inherent in an active site (amino
acids are chiral compounds)
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Week-9 CHEM113

– 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 PROPERTIES OF ALLOSTERIC ENZYMES

• Enzyme Inhibitor: a substance that slows down or • All allosteric enzymes have quaternary structure:
stops the normal catalytic function of an enzyme by – Composed of two or more protein subunits
binding to it. • Have at least two binding sites:
• Competitive Inhibitors: Compete with the substrate for – Substrate and regulator binding site
the same active site • Active and regulatory binding sites are distinct from
– Will have similar charge & shape each other:
– Noncompetitive Inhibitors: Do not compete – Located independent of each other
with the substrate for the same active site – Shapes of the sites (electronic geometry)
– Binds to the enzyme at a location other are different
than active site • Binding of molecules at the regulatory site causes
changes in the overall three-dimensional structure of
REVERSIBLE COMPETITIVE INHIBITION the enzyme:
• A competitive enzyme inhibitor: resembles an enzyme – Change in three-dimensional structure of
substrate in shape and charge the enzyme leads to change in enzyme
• Binds reversibly to an enzyme active site and the activity
inhibitor remains unchanged (no reaction occurs) – Some regulators increase enzyme activity –
• The enzyme - inhibitor complex formation is via weak activators
interactions (hydrogen bonds, etc.). – Some regulators decrease enzyme activity -
• Competitive inhibition can be reduced by simply inhibitors
increasing the concentration of the substrate.
FEEDBACK CONTROL
REVERSIBLE NONCOMPETITIVE INHIBITION
• A noncompetitive enzyme inhibitor decreases enzyme • Feedback Control: A process in which activation or
activity by binding to a site on an enzyme other than inhibition of the first reaction in a reaction sequence
the active site. is controlled by a product of the reaction sequence.
• Causes a change in the structure of the enzyme and • Regulators of a particular allosteric enzyme may be:
prevents enzyme activity. – Products of entirely different pathways of
• Increasing the concentration of substrate does not reaction within the cell
completely overcome inhibition. – compounds produced outside the cell
• Examples: Heavy metal ions Pb2+, Ag+, and Hg2+. (hormones)

IRREVERSIBLE INHIBITION PROTEOLYTIC ENZYMES AND ZYMOGENS

• An irreversible enzyme inhibitor inactivates enzymes • 2nd mechanism of regulating enzyme activity:
by forming a strong covalent bond with the enzyme’s – Production of enzymes in an inactive form
active site. (zymogens)
– The structure is not similar to enzyme’s – Zymogens are “turned on” at the
normal substrate appropriate time and place. It is the
– The inhibitor bonds strongly and increasing inactive precursor of a proteolytic enzyme.
substrate concentration does not reverse – Example: proteolytic enzymes: Most
the inhibition process digestive and blood-clotting enzymes are
– Enzyme is permanently inactivated. proteolytic enzymes
– E.g., Chemical warfare agents (nerve gases) – Hydrolyze peptide bonds in proteins
and organophosphate insecticides – Proteolytic enzymes are generated in an
inactive form and then converted to their
REGULATION OF ENZYME ACTIVITY active form

• 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
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COVALENT MODIFICATION OF ENZYMES PENICILLIN

• 3rd Mechanism for regulation of enzyme activity • Accidently discovered by Alexander Fleming in 1928
• Covalent modification: A process in which enzyme • Several naturally occurring penicillin and numerous
activity is altered by covalently modifying the synthetic derivatives have been produced
structure of the enzyme: • All have structures containing a four-membered Beta-
– Involves adding or removing a group from lactam ring fused with a five-membered thiazolidine
an enzyme ring
• Most common covalent modification: addition and • Selectively inhibits transpeptidase by covalent
removal of phosphate group: modification of serine residue
– Phosphate group is often derived from an • Transpeptidase catalyzes the formation of peptide
ATP molecule. cross links between polysaccharides strands in
– Addition of the phosphate bacterial cell walls
(phosphorylation) catalyzed by a Kinase
enzyme CIPRO
– Removal of the phosphate group • The antibiotic ciprofloxacin hydrochloride (Cipro for
(dephosphorylation) catalyzed by a short)
phosphatase enzyme. • Considered the best broad-spectrum antibiotics
– Phosphate group is added to (or removed because it is effective against skin and bone infections
from) the R group of a serine, tyrosine, or as well as against infections involving the urinary,
threonine amino acid residue in the enzyme gastrointestinal, and respiratory systems
regulated. • It is the drug of choice for treatment of traveler’s
diarrhea
• Bacteria are slow to acquire resistance to Cipro.
PRESCRIPTION DRUGS THAT INHIBIT ENZYME ACTIVITY – Biochemical threats associated with
• ACE ( angiotensin-converting enzymes) inhibitors terrorism has thrust Cipro into the spotlight
– an octapeptide hormone involved in blood because it is effective against anthrax.
pressure regulation. Increases blood
pressure by narrowing blood vessel. MEDICAL USES OF ENZYMES
– present as zymogen in the form of • Diagnose certain diseases:
angiotensinogen. – Enzymes produced in certain organ/tissues
example: Lisinopril if found in blood may indicate certain
damage to that organ/tissue
ANTIBIOTICS THAT INHIBIT ENZYME ACTIVITY Enzyme Conditions Indicated by
• An antibiotics is a substance that kills bacteria or Abnormal level
inhibits their growth Lactate Heart disease, liver disease
• Antibiotics usually inhibit specific enzymes dehydrogenase (LDH)
essential to life processes of bacteria Creatinine Heart disease
• Two families of antibiotics considered in this phosphokinase (CPK)
discussion are sulfa drugs and penicillin Aspartate Heart disease, liver disease,
aminotransferase (AST) muscle damage
SULFA DRUGS Alanine Heart disease, liver disease,
• Many derivatives of sulfanilamide collectively called aminotransferase (ALT) muscle damage
sulfa drugs exhibit antibiotic activities Gamma-glutamyl Heart disease, liver disease
• Sulfanilamide is structurally similar to PABA (p- transpeptidase (GGPT)
aminobenzoic acid) Alkaline phosphatase Bone disease, liver disease
• 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