BIOCHEMISTRY (LECTURE) ENZYME: COFACTOR

4. ENZYMES Small Organic Molecules Inorganic Ion

Derived from dietary Derived from dietary
Overview
vitamins minerals
- Proteins that act as a (true) catalyst for Also called as co-enzymes or
Nonmetallic ion cofactor: Cl-
biochemical reactions. co-substances
o TRUE CATALYST
 Sasama siya mismo sa
reaction then papabilisin nya.
Pag natapos na reaction
(reactant > product), andun pa
rin yung enzyme. Hindi sya
nacoconvert into something
else. It remains unchanged.
 Biological function of an
enzyme.
- Undergo all the reactions of proteins including
denaturation.
- EXAMPLE: Chymotrypsin
o Has an almost spherical shape so
considered to be a “globular protein”.

For Coenzyme or Organic

- There are thousands of enzymes inside a

person’s body.
ENZYMES: CLASSES
Simple Enzyme
- Protein only (amino acid chains)
Conjugated Enzyme
- Non-protein part + protein part
ENZYMES: STRUCTURE
Apoenzyme
- Protein part of conjugated enzyme.
Cofactor
- Non-protein part of conjugated enzyme. - Important to take vitamins because kailangan
yun for the proper functioning of an enzyme.
Holoenzyme
For Inorganic
- Biochemically active conjugated enzyme.

NOTE: Substrate is separate from conjugated enzyme.

It is not part of conjugated enzyme.
Substrate ENZYMES: SIX MAJOR CLASSES
- Reactant in an enzyme-catalyzed reaction.
- Substance upon which the enzyme “acts”.
o Cinoconvert into products.
- (Fits inside your enzyme.)
- (Once na pumasok na sa loob ng enzyme yung
substrate, kung ano mang compound yun, yun
yung macoconvert na into products.)
- (Pag lumabas na yung substrate, hindi na sya
substrate, nag-iba na siya ng structure so ang
tawag na sa kanya ay product.)

Ways to Determine the Class of a Reaction

1. Determine the substrate (left side).
a. The one wherein the structure is
expanded.
2. Reactant vs. Product
a. Compare the top-most carbon
(unchanged or not)
b. Bottom C
c. Middle C
3. Titignan saang subclass/class ng enzyme
GENERAL EQUATION FOR CHEMICAL REACTION: belong yung reaction or changes.
Reactants (Substrate) → Product/s ENZYME CLASS 1: OXIDOREDUCTASES
ENZYME: NOMENCLATURE
Non-Systematic Way of Naming Enzymes
- Suffix –ase identifies it as an enzyme
o Examples:
 Urease
 Sucrase
 Lipase
- EXCEPTION: the suffix –in is still found in the
names of some digestive enzymes.
o Examples:
 Trypsin ENZYME CLASS 2: TRANSFERASES
 Chymotrypsin
 Pepsin
- Type of reaction
o Oxidase: Catalyzes an oxidation rxn.
o Hydrolase: Catalyzes a hydrolysis rxn.
- Identity of substrate
o Glucose oxidase*
o Ptruvate carboxylase
o Succinate dehydrogenase
- General nature of substrate
o Lipase
 (Substrate happens to be in
LIPIDS.) ENZYME CLASS 3: HYDROLASES
o (Protease → PROTEINS.)
Systematic Way of Naming (following rules)
- Example: ATP: creatine phosphotransferase
- (EC 2.7.3.2)
o Transferases (EC 2)
o Transferring phosphorous-containing
groups. (EC 2.7)
o Phosphotransferases with a
nitrogenous group as acceptor. (EC
2.7.3)
o Creatine kinase. (EC 2.7.3.2)
- Divided into 6 major classes.
- Developed by IUBMB
o International Union of Biochemistry
and Molecular Biology.
ENZYME CLASS 4: LYASES Fischer Mechanism
- Proposed by Fischer.
- Also known as the “Lock-and-Key Model”.
- (BEFORE BINDING: Part ng substrate na
kakabit sa enzyme ay saktong-sakto – parang
jigsaw puzzle.)
- (AFTER BINDING: Since perfect match ang
substrate at active site ng enzyme, edi sakto
lang.)
Koshland Mechanism
- Proposed by Koshland.
- “Induced-Fit Model”
- (BEFORE BINDING: Part ng substrate na
ENZYME CLASS 5: ISOMERASE magbabind sa active site and yung mismong
active site ay hindi parehas, walang
correspondence, hindi perfect fit.)
- (AFTER BINDING: Once nag-enter na yung
substrate sa active site ng enzyme, the active
site of an enzyme will undergo some changes
in such a way that it will accommodate the
shape of your substrate.)
- (Binding of a substrate will induce the active
site of the enzyme in such a way that your
substrate will have a fit.)
- MAS ACCEPTABLE NA EXPLANATION
o Because there are enzymes that have
ENZYME CLASS 6: LIGASES more than 1 substrate.

MODELS OF ENZYME ACTION

Active Site ENZYME PROPERTIES: ACTIVE SITE
- Place where substrate binds to enzyme. - An asymmetric pocket wherein biological
- Some enzymes have more than 1 active site. reactions are catalyzed.
- Contains amino acid side chains that create a
Enzyme Substrate Complex 3-dimensional surface complementary to the
- Intermediate reaction species formed when substrate.
substrate binds with the active site.

2 MODELS FOR SUBSTRATE BINDING TO ENZYME

- These will explain to you how scientists
understand the binding of a substrate to the
enzyme.
- MODELS: Tells how they describe the process.
ENZYME PROPERTIES: CATALYTIC EFFICIENCY
- Each enzyme molecule:
o Transforms 100-1000 substrate
molecules into product each second.
- It’s 10³-10⁸ times faster than uncatalyzed
reactions.
-
- EFFICIENCY: gano kasipag ang enzyme.
- Quantity ng End Product: CRUCIAL
o Pag marami na yung amount ng EP, it
will signal the system na yung pathway
na yun must be shut off (titigil na sa
paggawa ng EP).
o WHY? Kasi dumadami na sila eh hindi
naman kailangan ng ganon karaming
amount yung katawan.
o One way to stop the metabolic pathway
- Turnover number to produce EP further is when EP na
o The number of substrate molecules an mismo ang magbabind sa unang
enzyme molecule can process per enzyme in such a way na kapag
second when saturated with the nagbind sya dun, your E1 will no longer
substrate. be able to bind your initial substrate.
o Equivalent to the catalytic rate ENZYME PROPERTIES: LOCATION WITHIN THE CELL
constant (kcat).
 Cat = catalysis - Many enzymes are localized in specific
 k = equilibrium constant organelles within the cell (compartmentalized).
o (Kunyari ang enzyme ay punong-puno
ng substrate > mabilis ang trabaho
non, kasi after maconvert ang isang REACTION VELOCITY (V)
substrate into product, di na mag-iintay
ng another substrate kasi saturated - Number of substrate molecules converted to
nga eh so meron na kagad available > product per unit time.
tatrabho na agad enzyme.) - Expressed as μmol of product formed per
minute.
o μ = micromol
o mmol = millimol
- (VELOCITY = bilis)
o (Gaano kabilis o katulin ang reaction
catalyzed by an enzyme.)
- (Gaano kadaming substrate yung naconvert ng
enzyme into a product per unit time.)
Factors that Affect Reaction Velocity
- Temperature
- pH
- Substrate Concentration
- Presence of Inhibitors
- Presence of Cofactors
ENZYME PROPERTIES: SPECIFITY FACTOR: TEMPERATURE
- Enzymes are highly specific. - Optimum temperature for human enzymes is
o Interacts with 1 or few substrates. 37°C (body temperature).
o Catalyzes only 1 type of chemical - Increased temperature (high fever) leads to
reaction. decreased enzyme activity.
ENZYME PROPERTIES: COFACTORS
- Non-protein portion of the enzyme that is
needed for enzymic activity.
ENZYME PROPERTIES: REGULATION
- Enzymes can be activated or inhibited so that
the rate of product formation responds to the
need of the cell.
o ACTIVATED: gumagawa ng product.
o INHIBITED: stop
 - (Kung anong factor inoobserve, yun yung Turnover Number
ilalagay sa x-axis ng graph.)
- (GRAPH: Inverted parabola) - Number of substrate molecules converted to
- (Tumataas temp = umiinit due to vibrations of product per second per enzyme molecule
molecules.) under conditions of optimum temperature and
- (Optimum temp = pinakamabilis na rate ng pH.
enzyme na makapagcatalyze ng reaction.)
FACTOR: pH
- Most enzymes have optimal activity in the pH
range of 7.0-7.5
- EXCEPTION:
o Pepsin: Optimum pH = 2.0
o Trypsin: Optimum pH = 8.0

- (Values on the right = pinakamataas na

number ng substrates na kayang i-catalyze ng
bawat enzyme kung ang temp at pH ay
optimum.)
- (Optimal activity = Reaction rate)
- (As pH increases, some of the side chains on FACTOR: ENZYME CONCENTRATION
the polypeptide chain of enzymes become
protonated while others become deprotonated; - Trend:
depends on the value of pH.) o At a constant substrate concentration,
o (Side chains are the reason kung bakit enzyme activity ↑ as enzyme
siya na-fold ng ganon in the first place.) concentration ↑
- (Mag-iiba na yung itsura ng enzyme;
madedenature na siya > hindi na magiging
functional para mag-catalyze ng reaction.)
FACTOR: SUBSTRATE CONCENTRATION
- Trend: ↑ substrate concentration, ↑ enzyme
activity (constant enzyme concentration)
- Substrate saturation
o Concentration at which it reaches its
maximum rate.
o All of the active sites are full. - (How “directly proportional” graph looks like.)
- (Limiting factor: Active Site – found in an
enzyme; where catalytic process occurs.)
- (Enzyme increases > more active site will
accommodate substrates > faster reaction >
more products will be formed.)
FACTOR: COFACTOR
- Coenzymes
- Metal Cofactors
- Small organic molecules which help and
facilitate enzyme reaction.

- (Concentration = dami)
- (Saturated = sagad)
- (Pag inextend yung graph, it will flat out forming
a plateau.)
- (Plateau part = lahat ng enzyme ay fully-
booked.)
- (Kahit magdagdag nang magdagdag ng
substrate, maghihintay pa rin siya ng turn niya
na iaccommodate sya ng active site ng enzyme
> maintained reaction rate = hindi bababa kasi
wala namang nabawas sa enzyme; lahat fully
functional, fully-booked nga lang.)
 - May be any of the following: HOW ENZYMES WORK: ENERGY CHANGES
o Part of the prosthetic group. OCCURRING DURING THE REACTION (cont.)
o May act independently facilitating
enzyme reactions. - Trend: ↓ free energy of activation, ↑ molecules
that have sufficient energy to pass through the
transition state, ↑ rate of reaction.

- (One of the things that is affected by drinking

vitamins is the proper functioning of enzymes.)
FACTOR: INHIBITORS
- Any substance that can diminish the velocity of
an enzyme-catalyzed reaction. - (Transition State: Highest part of the graph;
o May chemical structure. peak.)
- (Papabagalin or totally blocked the reaction.) o Your reactants are in the middle of
being products.
HOW ENZYMES WORK: ENERGY CHANGES o They do not look like exactly like the
OCCURRING DURING THE REACTION initial reactant but they are not
products yet.
Free Energy of Activation - (W/ enzyme: makes the conversion of the
- Energy difference between that of the reactants into products faster.)
reactants and a high-energy intermediate that o REASON: Kaya niyang pababain TS in
occurs during the formation of product. such a way na mas maraming
reactants makakatawid para ma-
Rate of Reaction convert into products.
o Enzyme napapababa activation energy.
- Determined by the number of energized
molecules. KINETICS OF ENZYME CATALYSIS: MICHAELIS-MENTEN
EQUATION

k₁ k₂
E+S↔
k₋₁ ES →E+P
o E = (free) Enzyme
o S = Substrate
o ES = Enzyme-Substrate Complex
o P = Product
- It describes how reaction velocity varies with
substrate concentration.
- EQUATION:
𝑉ₘₐₓ [𝑆]
- (x-axis: Reaction path: It will tell u kung saang 𝑉ᵢₙᵢₜᵢₐₗ
𝐾ₘ + [𝑆]
mga substances nagsimula yung reaction then
in the end, maiiba siya.) o V = Velocity
- (y-axis: Energy: Mas mataas o mas malaki, mas o S = Substrate
mahirap macrossover.) o Kₘ = Michaelis-Menten Equation
- (May part na parehas reaction w/ catalyst and o [ ] = Concentration in Molarity
w/o catalyst – EXCEPT the middle part) Important Conclusions About Michaelis-Menten
o Means: For the reactant and product Kinetics
side, parehas sila ng energy
requirement. - Kₘ = ½ Vₘₐₓ
o Same start, same ending o Small Kₘ (closer to 0)
- (Pag mas mababa yung elevated part sa gitna,  Enzyme has a high affinity for
mas maraming reactants na pwedeng mag- substrate.
cross para ma-convert into products.) o Large Kₘ
- (Mas mataas elevated part = no product, or  Enzyme has a low affinity for
very dew, or very slow process before substrate.
makagawa ng product.)
 - DERIVATION:

- Enzyme 1: Higher affinity for the substrate. o It was arranged in such a way that the
ending will look like a slope-intercept
ORDER OF REACTION form.
Zero Order - Intercept on the y-axis: 1/Vmax
- Intercept on the x-axis: -1/Km
- [S] >>> Km - X-axis: 1/[S]
o (much greater than) - Y-axis: 1/V₀
- The velocity is constant and equal to Vmax.
- (Sobrang dami na ng substrate, velocity of
reaction becomes constant.)
First Order
- [S] <<< Km
o (much less than)
- The velocity is approximately proportional to
the substrate concentration.
- (Sobrang liit ng concentration ng substrate sa
beginning; reaction rate proportional sa - 2 ways of mathematically expressing the
substrate concentration.) catalysis reaction that happens to your enzyme
(Michaelis-Menten and Lineweaver-Burke).
ENZYME INHIBITION
Enzyme Inhibitor
- A substance that slows down or stops the
normal catalytic function of an enzyme by
binding to it.
Modes of Inhibition
- Irreversible
- Reversible
o Competitive
o Noncompetitive
o Uncompetitive
LINEWEAVER-BURKE PLOT
ENZYME INHIBITION: IRREVERSIBLE
- Double-reciprocal plot.
- FUNCTIONS: Irreversible Enzyme Inhibitor
o To calculate Km and Vmax. - Inactivates enzymes by forming a strong
 Cinocompute most especially covalent bond with the enzyme’s active site.
kapag nagdadagdag na ng - The structure is not similar to enzyme’s normal
inhibitor; kapag minemeasure substrate.
na yung effect ng inhibitor.
Need kasi may iba’t-ibang
klase ng inhibitor.
o To determine the mechanism of action
of enzyme inhibitors.
- EQUATION:
1 𝐾ₘ 1 1
= ( )+
𝑉 𝑉ₘₐₓ [𝑆] 𝑉ₘₐₓ

y= m x + b
o Slope = Michaelis-Menten constant
and maximum velocity
o Intercept = 1/Vmax = reciprocal of the
maximum velocity
- - (Point of no return.)
 - (Once the enzyme was blocked by an inhibitor, ENZYME INHIBITION: COMPETITIVE
its catalytic process will be totally disrupted – it
will no longer catalyze.)
- (Pag nagform ng covalent bond – wherein the
electrons are shared – hindi yun basta-basta
natitibag.)
- (Once formed, the structure of an enzyme will
change.)
IRREVERSIBLE INHIBITION: PENICILLIN
- Suicide inhibition closely resembles
competitive inhibition. - (Figure 1: Substrate binds to an enzyme
through its active site > forms the enzyme-
substrate complex > produce a product and
release again the free enzyme.)
- (Figure 2: Instead of a substrate, a competitive
inhibitor will be the one that will bind to an
enzyme’s active site > forming an enzyme
inhibitor complex > walang product.)
- WHY is it called as COMPETITIVE INHIBITOR?
o Meron siyang part sa structure niya na
COMPLETELY SAME sa substrate and
yun yung gagamitin niya para mag-bind
sa active site > hindi irereject ng AS.
o Makikipag-unahan ang inhibitor na ‘to
papuntang AS ng enzyme (competing
against your substrate for the AS).
- EXAMPLES: Atorvastatin and Simvastatin
METABOLIC PATHWAY

- (Steps 1-3: Depicts how the bacterial cell wall

is being made.)
o (Protein will catalyze the creation of
peptidoglycan layer – composition of
bacterial cell wall)
o Pentaglycine chain: the one in yellow
o (In the presence of binding protein, pag
nag-attach sa yellow, it will catalyze the
formation of the bridge between the
two vertical polypeptide chain.)
- (Steps 4 & 5)
o Penicillin will bind to the binding
protein (mismong enzyme na
nagcacatalyze ng reaction na
panggawa ng peptidoglycan layer.) >
then hindi na siya matatanggal >
madidisrupt na yung paggawa ng cell
wall; di na niya masysynthesize
paggawa ng peptidoglycan layer.
INHIBITOR OF ENZYME ACTIVITY: REVERSIBLE
- May or may not alter the conformation of the
enzymes.
- When it binds to an enzyme…
o It alters enzyme activities - Known as the “Synthesis of the Cholesterol”
o It’s not permanent and can be - (First Committed Step: Very crucial and very
disrupted by the following: important because once the Mevalonic Acid is
 Presence of excess substrates produced, itong buong reaction na to, good to
 Presence of substrates with go na; dire-diretso na; wala na hahadlang
higher affinity for the enzyme hanggang ma-form ang ultimate product.)
(than the inhibitor). - (ULTIMATE PRODUCT: Cholesterol)
- (STATIN: nagawang gamot to inhibit the first
committed step.)
SOME SUBSTANCES THAT BELONG TO STATIN FAMILY o Noncompetitive Inhibitor of this
reaction: Heavy Metals
- (SIMILARITY sa COMPETITIVE: Path 1)
- (SIMILARITY sa UNCOMPE: Path 2)
ENZYME INHIBITION: UNCOMPETITIVE

- They are the synthesized substances which will

act as the competitive inhibitor for the
substrate (HMG-CoA).
- Once the statin binds to that substrate, the
reaction will be temporarily disrupted > walang
Mevalonic Acid na mafoform > wala ring
Cholesterol > bababa yung cholesterol in the
body.
- Since these are inhibitors, there are parts in
their structures that are same with HMG-CoA.
- (EQUATION 1 and FIG 1: Normal Catalytic
ENZYME INHIBITION: NON-COMPETITIVE Process.)
- (Presence of Uncompetitive Inhibitor: It will
follow the same pathway until the formation of
enzyme-substrate complex. Once it is formed,
the uncompetitive inhibitor will bind to that
complex > forming another complex called
“enzyme substrate inhibitor complex”.)
- (Sa enzyme substrate complex lang siya
magbabind, wala nang iba. So iintayin ny
amuna ma-form yun before siya mag-bind = NO
PRODUCT.)
- EXAMPLE: Glyphosate (herbicide)
o Beneficial to plants because this
removes the weeds.
o SHIKIMATE PATHWAY: Metabolic
pathway (in plants) wherein this
specifically acts upon.
- Opposite end of Competitive Inhibitor.
- It will not compete against the substrate ULTIMATE TEST TO DIFFERENTIATE WHICH TYPE OF
because it will bind to your enzyme in locations IHNIBITION HAS TAKE PLACE: through graphing.
other than the active site.
- (PATH 1: NI will bind to a free enzyme >
producing an enzyme-inhibitor complex +
substrate > producing another complex called
“enzyme-substrate-inhibitor complex” =
walang product)
- (PATH 2: Substrate will bind to a free enzyme >
forming enzyme-substrate complex +
noncompetitive inhibitor > forming “enzyme
substrate inhibitor complex” = NO product
formed.)
- (Fig 1: Fit na fit substrate sa AS = Product)
- (Fig 2: naka-enter ang substrate sa AS pero
hindi nakapag-bind effectively with the
presence of NI; maluwag = nagbago ang
conformation ng enzyme when a NI binds to it
kaya kahit na nakapasok yung substrate, wala
pa ring product na mafoform.)
- EXAMPLE: Ferrochelatase (enzyme)
o Catalyzes the binding or attachment of
Fe2+ to your porphyrin = heme (combi)
 Heme: nagdadala ng oxygen to
the different parts of the body.