CHEM123

Ms. Rose Dyane F. Nunag, RMT, MPH

S.Y 2023- 2024: MIDTERMS

WEEK 9: ENZYMES & KREB’S CYCLE NOMENCLATURE & CLASSIFICATION OF

ENZYME ENZYMES
● It is a compound usually a protein, that acts Named about the function of the enzyme, type
as a catalyst for a biochemical reaction of reaction catalyzed and the substrate identity
● Cause cellular reaction to occur millions of 1. Suffix most enzymes end in the suffix “ase”
times faster - Ex. Urease, Sucrase, Lipase
● Not consumed during the reaction but - Exception: digestive enzymes
merely help the reaction occur more rapidly - Ex. Trypsin, chymotrypsin, pepsin
● Mostly, are globular proteins 2. Type of reaction catalyzed by an enzyme is
● Medical Uses of Enzymes often used as a prefix
- Used to diagnose and to monitor - Ex. Oxidase- oxidation reaction
treatment of certain diseases - Hydrolase- hydrolysis reaction
- Appearance of these enzymes in the 3. Identity of substrate and type of reaction
blood often indicates that there is catalyzed
tissue damage in an organ and that - Ex. Glucose oxidase, pyruvate
cellular contents are spilling out into carboxylase, succinate
the bloodstream dehydrogenase
ENZYME STRUCTURE: Substrate
• Simple enzyme - Reactant in an enzyme – catalyzed
- Composed only of protein reaction.
• Conjugated Enzyme
- Has a non – protein part in addition to a ENZYME CLASSIFICATION
protein part.
- Apoenzyme- Protein of the conjugated
enzyme
- Cofactor- Non – protein part of the
conjugated enzyme
- Holoenzyme- Biochemically active
conjugated enzyme produced from an
apoenzyme and a cofactor
- Combined apoenzyme and cofactor entity
- Coenzyme- Serves as a cofactor in a
conjugated enzyme

MODELS OF ENZYME ACTION

Enzyme active site
- Small part of an enzyme's structure that is
actually involved in catalysis.

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 - A three – dimensional entity formed by ENZYME SPECIFICITY
groups that come from different parts of the - Extent to which an enzyme’s activity is
protein chains restricted to a specific substrate, a specific
Enzyme- Substrate Complex group of substrate, a specific type of
- The intermediate reaction species that is chemical bond, or a specific type of
formed when a substrate binds to the chemical reaction.
active site of an enzyme. - Degree of specificity is determined by the
active site.

1. Absolute Specificity
- Catalyze only one reaction
- Most restrictive of all specificities
- Catalase – enzyme with absolute
specificity
2. Group Specificity
LOCK- AND- MODEL - Act only on molecules that have a
- Active site in the enzyme has the fixed, specific functional group, such as
rigid geometrical conformation. hydroxyl, amino or phosphate
- Substrate with a complementary geometry groups.
can be accommodated. - Carboxyl-peptidase is group
specific.
3. Linkage Specificity
- Act on the particular type of bond,
irrespective to the rest of the
molecular structure.
- Phosphatases hydrolyze phosphate
– ester bonds in all types of
phosphate esters
- Most general of the common
species
INDUCED- FIT MODEL 4. Stereochemical specificity
- Enzyme’s active site is not rigid and static. - Act on a particular isomer
- There’s a constant change in shape
- Allows for changes in the shape or FACTORS THAT AFFECT ENZYME ACTIVITY
geometry of the active site of an ● Enzyme Activity
- enzyme to accommodate a substrate. - Measures the rate at which an
- Result of the enzyme’s flexibility; it adapts enzyme converts substrate to
the incoming substrate. products in a biochemical reaction

● Factors that affects enzyme activity

- Temperature
- pH
- Substrate Concentration
- Enzyme Concentration

1. TEMPERATURE
● Measure of kinetic energy of molecules.
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 ● Higher temperatures mean molecules are 3. SUBSTRATE CONCENTRATION
moving faster and colliding more frequently ● Increased concentration of substrate will
● Optimum temperature obtain the enzyme activity.
- Temperature at which an enzyme ● Turnover number
exhibits maximum activity - Number of substrate molecules
transformed per minute by one
molecule of enzyme under optimum
conditions of temperature, pH and
saturation.
4. ENZYME CONCENTRATION
● Kept in a low number because enzymes
are not consumed in the reaction.
● The greater the enzyme concentration, the
greater the reaction rate.

2. pH
● the charge on acidic and basic
amino acids located at the active
site depends on pH
● small pH changes can result in
enzyme denaturation and
subsequent loss of catalytic activity.
● Biochemical buffers help maintain
the optimum pH for an enzyme.
● Can also affect substrate, causing ENZYME INHIBITION
either protonation or deprotonation ● Enzyme Inhibitor
of groups on the substrate. - Substance that slows or stops the
● Optimum pH normal catalytic function of an
- pH at which an enzyme enzyme by binding to it.
exhibits maximum activity 1. Reversible Competitive Inhibition
- physiological pH ranges from 2. Reversible Non competitive inhibition
7.0 – 7.5 3. Irreversible inhibition
- Pepsin- Active in the
stomach, functions best at pH TYPES OF ENZYME INHIBITION
2.0 REVERSIBLE COMPETITIVE INHIBITION
- Trypsin- Operates in the - Molecule that sufficiently resembles an
small intestines, function best enzyme substrate in shape and charge
at pH 8.0 distribution that it can compete with the
substrate for occupancy of the enzymes
active site
- Remains in changed as it binds to the
enzyme's active site.
- Reversible process because it is
maintained but weak interactions
- Can be reduced by increasing the
concentration of the substrate.
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REVERSIBLE NON- COMPETITIVE INHIBITION ENZYME WITH TWO OR MORE PROTEIN
- Molecule that decreases enzyme activity CHAINS & 2 KINDS OF BINDING SITES
by binding to a site on an enzyme other Regulators
than the active site. - Substance that bind at the regulatory sites
- Presence of this causes a change in the of allosteric enzymes
structure of the enzyme sufficient to 1. Positive Regulator
prevent the catalytic groups at the active - Increase enzyme activity
site from properly affecting their catalyzing - The shape of the active site is
action. changed such that it can more
IRREVERSIBLE INHIBITION readily accept substrates.
- Molecule that inactivates enzymes by 2. Negative Regulator
forming a strong covalent bond to an amino - Decrease enzyme activity
acid side – chain group at the enzymes - Changes to the active site are such
active site. that substrate is less readily
- Do not have structures similar to that of the accepted.
enzyme’s normal substrate.
FEEDBACK CONTROL
REGULATION OF ENZYME ACTIVITY - A process in which activation or inhibition
1. A cell that continually produces a large of the first reaction in a reaction sequence
amount of enzyme for which substrate is controlled by a product of reaction
concentration is always very low is wasting sequence.
energy. The production of the enzyme - Negative Feedback
needs to be “ turned off”.
PROTEOLYTIC ENZYMES & ZYMOGENS
2. A product of an enzyme- Catalyzed ● Proteolytic Enzymes
reaction that is present in plentiful amounts - Catalyzes the breaking of peptide
in a cell is a waste of energy if the enzyme bonds that maintain the primary
continues to catalyze the reaction that structure of protein.
produces the product. The enzyme needs - Generated in an inactive form and
to be turned off. converted to active form when they
are needed.
ALLOSTERIC ENZYME
1. Have Quaternary Structure; (2 or more ● Zymogen
protein chains). - Inactive precursor of a proteolytic
2. Have 2 kinds of binding sites (for enzyme.
substrates and for regulators). - Activation of a zymogen requires an
3. Active and regulatory binding sites are enzyme – controlled reaction that
distinct from each other in both location moves some part of the zymogen
and shape. structure which changes the 3 –
4. Binding of a molecule at the regulatory site dimensional structure of zymogen,
causes changes in the overall three which affects active site
dimensional structure of the enzyme, conformation.
including structural changes at the active
site.

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COVALENT MODIFICATION OF ENZYME STAGE 2- ACETYL GROUP FORMATION
● Process in which enzyme activity is altered ● The small molecules from Stage 1 are
by covalently modifying the structure of the further oxidized.
enzyme through attachment of a chemical ● End products of these oxidations is acetyl
group or removal of a chemical group from CoA
a particular amino acid within the enzyme ● Involves numerous reactions:
structure. - Reactions occur both in cytosol
● Phosphorylation (glucose metabolism) as well as
- Process of addition of the phosphate mitochondria (fatty acid metabolism)
group to the enzyme by protein of the cells.
kinases
● Dephosphorylation STAGE 3- CITRIC ACID CYCLE
- Removal of the phosphate group ● Takes place in inside the mitochondria
from the enzyme by phosphatases. ● First intermediate of the cycle is citric acid
therefore designated as Citric acid cycle
OVERVIEW OF THE BIOCHEMICAL ENERGY ● In this stage acetyl group is oxidized to
PRODUCTION produce CO2 and energy
● Energy needed to run human body is ● The carbon oxide we exhale comes
obtained from food primarily from this stage
● Multi-step process that involves several ● Most energy is trapped in reduced
different catabolic pathways coenzymes NADH and FADH2
● There are four general stages in the ● Some energy produced in this stage is lost
biochemical energy production process: in the form of heat
- Stage 1: Digestion
- Stage 2: Acetyl group formation, STAGE 4- ELECTRON TRANSPORT CHAIN &
- Stage 3: Citric acid cycle OXIDATIVE PHOSPHORYLATION
- Stage 4: Electron transport chain ● Takes place in mitochondria
and oxidative phosphorylation, ● NADH and FADH2 are oxidized to release
H+ and electrons
● Each stage also involves numerous ● H+ are transported to the intermembrane
reactions space in mitochondria
STAGE 1- DIGESTION ● Electrons are transferred to O2 and O2 is
● Begins in mouth (saliva contains starch reduced to H2O
digesting enzymes), continues in the ● H+ ions reenter the mitochondrial matrix
stomach (gastric juice), completed in small and drive ATPsynthase reaction to produce
intestine: ATP
- Results in small molecules that can ● ATP is the primary energy carrier in
cross intestinal membrane into the metabolic pathways
blood ● Citric acid cycle: A series of biochemical
● End Products of digestion: reactions in which the acetyl portion of
- Glucose and monosaccharides from acetyl CoA is oxidized to carbon dioxide
carbohydrates and the reduced coenzymes FADH2 and
- Amino acids from proteins NADH are produced
- Fatty acids and glycerol from fats ● Also known as tricarboxylic acid cycle
and oils (TCA) or Krebs cycle:
● The digestion products are absorbed into - Citric acid is a tricarboxylic acid –
the blood and transported to body’s cells TCA cycle
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 - Named after Hans Krebs who ● When ATP levels are low ADP, ADP
elucidated this pathway activates citrate synthase
● Two important types of reactions: ● Similarly, ADP and NADH control isocitrate
- Oxidation of NAD+ and FAD to dehydrogenase:
produce NADH and FADH2 - NADH acts as an inhibitor
- Decarboxylation of citric acid to - ADP as an activator.
produce carbon dioxide ● The electron transport chain (ETC)
- The citric acid cycle also produces 2 facilitates the passage of electrons trapped
ATP by substrate level in FADH2 and NADH during citric cycle
phosphorylation from GTP ● ETC is a series of biochemical reactions in
● Summary of citric acid cycle reactions: which intermediate carriers (protein and
- Acetyl CoA + 3NAD + FAD + GDP + non-protein) aid the transfer of electrons
Pi + 2H2O 2CO2 + CoA-SH + and hydrogen ions from NADH and FADH2
3NADH + 2H+ + FADH2 + GTP ● The ultimately receiver of electrons is
Step 1: Formation of Citrate molecular oxygen
Step 2: Formation of Isocitrate ● The electron transport (respiratory chain)
Step 3: Oxidation of Isocitrate and Formation of gets its name from the fact electrons are
CO2: involves oxidation– reduction as well as transported to oxygen absorbed via
decarboxylation respiration
Step 4: Oxidation of Alpha- Ketoglutarate and ● The overall ETC reaction: 2 H+ + 2e- + 1/2
Formation of CO2 O2 H2O + energy
Step 5: Thioester bond cleavage in Succinyl CoA ● Energy is used to synthesize ATP in
and Phosphorylation of GDP oxidative phosphorylation
Step 6: Oxidation of Succinate ● Note that 2 hydrogen ions, 2 electrons, and
Step 7: Hydration of Fumarate one half-oxygen molecule react to form the
Step 8: Oxidation of L-Malate to Regenerate product water
Oxaloacetate ● This relatively straight forward reaction that
requires eight or more steps
● The reaction releases energy (exothermic
reaction)
● The energy released is coupled with the
formation of three ATP molecules per every
molecule of NADH processed through ETC
● The enzymes and electron carriers needed
for the ETC are located along inner
mitochondrial membrane
● They are organized into four distinct protein
complexes and two mobile carriers
● The four protein complexes tightly bound to
membrane:
● Complex 1: NADH-coenzyme Q reductase
REGULATION OF THE CITRIC ACID CYCLE ● Complex II: Succinate-coenzyme Q
● The rate at which the citric acid cycle reductase
operates is controlled by ATP and NADH ● Complex III: Coenzyme Q - cytochrome C
levels reductase
● When ATP supply is high, ATP inhibits ● Complex IV: Cytochrome C oxidase
citrate synthase (Step 1 of Citric acid cycle) ● Two mobile electron carriers are:
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 - Coenzyme Q and cytochrome c. - In the final stage of electron transfer, the
electrons from cyt a3, and hydrogen ion
COMPLEX 1: NADH- COENZYME Q (H+) combine with oxygen (O2) to form
REDUCTASE water
- NADH from citric acid cycle is the source of - O2 + 4H+ + 4e- 2 H2O
electrons for this complex - It is estimated that 95 % of the oxygen
- It contains >40 subunits including flavin used by cells serves as the final electron
mononucleotide (FMN) and several acceptor for the ETC
iron-sulfur protein clusters (FeSP)
- Net result: Facilitates transfer of electrons
from NADH to coenzyme Q
- Several intermediate reactions are involved
in this electron transfer

COMPLEX II: SUCCINATE- COENZYME Q

REDUCTASE
- Smaller than complex I
- Contains only four subunits including two
iron-sulfur protein clusters (FeSP)
- Succinate is converted to fumarate by this
complex
- In the process it generates FADH2
- CoQ is the final recipient of the electrons
from FADH2

COMPLEX III: COENZYME Q- CYTOCHROME-

c REDUCTASE
- Complex III contains 11 different subunits
- Several iron-sulfur proteins and
cytochromes are electron carriers in this
complex
- Cytochrome is a heme iron protein in which
reversible oxidation of an iron atom occurs
- Various cytochromes, e.g., cyt a, cyt b, cyt
c, differ from each other by:
- Their protein constituents
- The manner in which the heme is
bonded to the protein
- Attachments to the heme ring

COMPLEX IV: CYTOCHROME c OXIDASE

- Contains 13 subunits including two
cytochromes P.S. Watch the recorded discussion about enzymes & kreb’s
cycle for better understanding.
- The electrons flow from cyt c to cyt a to cyt
a3 Do not share without ACDN’s permission.

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