ALLOSTERIC SITE: Site other than the active site that may

CLINICAL CHEMISTRY
lead to either attachment of substrate to the enzyme’s active
- Activities, properties of enzymes in specimen to aid
site or inhibition of attachment
diagnosis and treatment
ISOENZYMES: different form of an enzyme with different
ENZYMES
genetic origins but catalyze the same reaction
- These are protein molecules that catalyzes a given
chemical reaction w/o being destroyed/altered
ISOFORMS: Results when an enzyme is subject to different
- Biological catalysts: causes reactions in the body to
post-transitional modification
take place
- Cellular catalysts: speed up rate of chemical
ENZYME STRUCTURES
reactions
a. Primary Structure: Refers to the sequence of amino
- catalysts: not consumed, destroyed, alter or
acids joined by peptide bonds to form a polypeptide
changed by the reaction
chain

CHARACTERISTICS
b. Secondary Structure: Conformation of the segments
- The reactions they catalyze are frequently reversible
of polypeptide chain
which means that they can synthesize and
- Made up of alpha helices or beta-pleated
decompose molecules
sheets which are maintained by hydrogen
- They are complicated types of protein in terms of both
bonds
structure and function
- They easily denatured with varying molecular weight
c. Tertiary Structure: Arises from the interactions
and mass
among side chains/groups of the polypeptide chain
- These enzymes are amphoteric which are capable of
ionizing either as acid or base
- Structure are bent and folded and maintained by
- They are synthesized in an inactive state and
covalent disulfide bond
operates in the presence of a cofactor
- Changes in enzyme concentration reflect changes in
d. Quaternary Structure: Separate bended & folded
state of health.
structures are put together to form a functional unit
- Operate at high rates
- Enzyme variants – LDH, Creatine kinase

COENZYMES - non-protein organic biochemical that takes

The enzyme action model
part in the enzyme reaction
a. Enzymes act through formation of enzyme substrate
- Essential to the catalytic activity as a
complex
CO-SUBSTRATE
b. The substrate must be bound to the active site of the
- Diffusible, heat stable, low molecular weight that when
enzyme
combined tightly to enzymes, the coenzyme will be
c. The enzyme-substrate complex will then break down
called Prosthetic group
to give the reaction products and free the enzyme
- E.g. NAD, Pyridoxal phosphate
d. All enzyme reactions are in theory reversible however,
in practice, reactions are usually more rapid in one
ACTIVATORS - Inorganic ionic cofactor (effector molecules)
direction than the other.
- increase the catalytic activity of an enzyme when it
binds to specific site
- Metabolic regulator of enzyme reaction Usually metal
Michaelis-Menten Curve/Plot: shows the relationship of the
ions (esp. divalent cations)
reaction velocity to the substrate concentration
- E.g. Mg++, Na+, K+, Zn++
● Km - a constant for specific enzyme and substrate
HOLOENZYME: the combined enzyme & coenzyme
under defined reaction conditions
- Expression of the relationship b/w velocity of
APOENZYME: Enzyme without a cofactor
an enzymatic reaction and substrate conc.
- half of the maximum velocity/speed of enzymes
PROSTHETIC GROUP: A coenzyme that cannot be removed
- amount of substrate needed for a particular enzymatic
from its attachment to an enzyme using dialysis
reaction
- E.g. Pyridoxal phosphate in transaminase reaction
V: measured velocity of reaction
SUBSTRATE: Substance acted upon by an enzyme & is
Vmax: max. velocity
converted into a new substance
S: substrate
Km: Michaelis-Mendel constant of enzyme for specific sub.
PRODUCT: Substance derived from a transformed substrate
Lineweaver-Burk Plot
ACTIVE SITE: Site where substrate interacts with enzymes
 - Transformation of Michaelis-Mendel curve - Catalyze reactions similar to the following: A > B
- Vmax: the reciprocal of the x-intercept of the straight - All reactions are reversible
line - Transformation from:
- Km: negative reciprocal of the x-intercept of the same - Cis (same side) to Trans (other)
line - L (OH on the left side) to D (OH grp position on
- More accurate and convenient determination right) form
- Aldehyde to Ketones (neither) ; contain carbonyl
ENZYME CLASSIFICATION AND NOMENCLATURE bond
- Based of catalytic activity - Ex: Glucose PO4 isomerase
- International Union of Biochemistry: Enzyme
Commission categorized enzymes into 6 classes f. Ligases
- Catalyzes joining of 2 substrate molecules coupled by
a. Oxidoreductase breaking of pyrophosphate bond
- Older names: Dehydrogenases & Oxidases - Enzymes causing bond formation between two
- Catalyze oxidation-reduction reactions molecule to form a larger molecule
- Catalyze the transfer of electrons from one molecule - Ex: Ligase, Animo Acyl t-RNA synthetase
(the oxidant) to another molecule (the reductant).
- Catalyze reactions similar to the following: A– + B → ENZYME NOMENCLATURE
A + B– where A is the oxidant and B is the reductant
- Assayed for investigation of cardiac & liver disorders TO CONSIDER:
- Ex: Glucose Oxidase, Cytochrome Oxidase, Lactate 1. Type of reaction catalyzed
dehydrogenase 2. Suffix “ASE” added to the name of the substrate
3. EMPIRICAL name
b. Transferases 4. Standard system (IUB and IUPAC)
- Catalyzes a group other than hydrogen
- Move an intact group of atoms (NH2 or PO4) from a. Systematic name
one molecule to another - Describes the nature of the reaction catalyzed
- Catalyze reactions similar to the following A + BX = - Numerical code designation prefixed w/ the letters
AX + B (ATP + Creatine = ADP+Creating PO4) E.C. (4 digit separated by decimal points)
- Gives information about liver damage - Example: E.C. 3.1.3.1 = ALP
- Ex: Aspartate Aminotransferase, Alanine and E.C. 3.1.3.2 = ACP
aminotransferase, Creatine kinase - 1st digit = denotes the class of the enzyme
- 2nd digit = sub-class of the enzyme
c. Hydrolases - 3rd digit = sub sub-class
- Catalyze the hydrolysis of various bonds - 4th digit = specific serial number
- Splits molecules with water as part of the reaction * This approach removes all ambiguity about the
process enzyme;’s identity
- Catalyze reactions similar to the following: L A + H20
=B+C b. Trivial name
- Three (3) Groups: - a.k.a Non- specific, Practical name, Working name
a. Esterases: ACP, ALP, Lipase - Uses acronyms and abbreviations
b. Peptidases: Leucine aminopeptidase, Pepsin - Examples: SGOT, SGPT
c. Glycosidases: Amylase,
Amylo-1,6-glucosidase

d. Lyases
- Removal of groups form substrate w/o hydrolysis
- Product: Double bond
- Responsible for splitting molecules or breaking of
bonds (C to C; C to O; C to N, etc.)
- Catalyze reactions similar to the following: A = B + C
- Assayed in disorders of skeletal muscles
- Ex: Aldolase, Glutamate decarboxylase, Pyruvate
decarboxylase

e. Isomerases
- Catalyze interconversion of geometric, optical or
positional isomers
- Responsible for the conversion of one isomer to
another
 ENZYME VARIANTS B. BILOCULAR ENZYME
- Found in both mitochondria and cell sap
- These are several distinct forms of enzymes
- Important diagnosis of specificity

A. ISOENZYMES: ENZYME KINETICS

- Multi-chained enzyme of similar activity - The general relationship among enzyme, substrate and
- Appear in specific organ, cell & cell organelle of product (E+S = ES = P+E)
similar organisms E: unchanged enzyme (catalysts)
- Ex: Lactate dehydrogenase- catalyzes S: substrate
interconversion of lactic and pyruvate acid P: product
- Widely distributed in the body: heart, liver, skeletal ES: postulated enzyme- substrate complex ; physical binding
muscle, kidney, erythrocyte (high act) ; lung, smooth to a substrate
muscle, brain (low act)
- Contains H (heart) & M (muscle) subunit
(polypeptide chains) = combine is 5 isoenzyme
fractions

Characteristics:
1. Electrophoretic mobility
2. Mobility in Ion Exchange Resin
3. Response to inhibition

A. FACTORS AFFECT BINDING OF AN ENZYME TO

4. Relative substrate specificities SUBSTRATE
Ex: ACP (RBC) less sensitive to a-naphthyl PO4 ● Energy
● Molecular compatibility
B. HETEROENZYMES ● Space availability
- Enzymes of similar catalytic activity but are specie ● Specificity
specific
B. THEORIES RELATED TO E-S COMBINATION
C. ALLOENZYME ● Lock and Key Theory
- Genetically transmitted enzyme - refers to the active site being complementary
- Important in defining biochemical characteristics of an in shape & size to the substrate (perfect)
individual - First presented by Emil Fisher

ORIGIN OF PLASMA ENZYMES Induced fit Model

- The enzyme changes in shape during binding to
Classification in the blood accommodate the substrate
A. PLASMA SPECIFIC ENZYMES - Theory maintains that the active site and the
- Generally secreted by liver substrate are, initially, not perfect matches for each
- Enzymes that insert their functions in plasma other.

B. NON-PLASMA SPECIFIC ENZYMES C. Factors that influence the enzymatic reaction

- No specific functions in plasma (they lack a. TIME – The rate of enzymatic reaction
activators/coenzyme) * If the catalytic activity of an enzyme on a substrate
- Two classes: is fast, this will mean a shorter reaction time
a. Enzymes of secretion: secreted in plasma at
high rate but rapidly disposed off to excretory b.SUBSTRATE CONCENTRATION: the rate at which
b. Enzymes associated w/ cellular metabolism: an enzymatic reaction proceeds and whether the
carry out their functions w/in the cells in forward or reverse reaction occurs
which they are formed - Direct relationship (more enzyme = steady)
- Michaelis-Menten: the substrate readily bind
Classification based on distribution to free enzymes at a low substrate concent
A. UNILOCULAR ENZYME TWO PHASES:
- Found only in one location (cell sap) - First order Kinetics
o Enzyme conc. is fixed; Substrate conc. is varied
o Rate of reaction is almost directly proportional to
 substrate conc. at low values
TYPES:
- Zero order Kinetics Reversible inhibition - Inhibitors are possible removed from
o When maximum velocity is reached, the rate of the system; enzyme is fully restored
increase in velocity is “O” - Physical processes that remove inhibitors: dialysis,
o Reaction rate is unaffected by increased substrate gel filtration
concentration Irreversible inhibition - Inhibitors covalently combine w/ the
o Dependent on enzyme concentration enzyme
- Physical methods are ineffective in separating
c. ENZYME CONCENTRATION: inhibitors from the enzymes
- Enzymes catalyzes physiologic reactions
and affects the rate EXAMPLES OF INHIBITORS:
- Direct relationship: The higher the enzyme 1. Excess substrate: causes competition b/w substrate
level the faster the reaction will proceed molecules for a single binding site

d. TEMPERATURE: 2. Product of reaction: maybe an inhibitor of forward reaction

- Increasing temp also increase reaction rate 3. E-S complex does not break to yield products
- Optimum temperature - °T considered 4. Chemical Substances
favorable for enzyme activity (30-37°C or 37
– 40°C) I. COENZYME CONCENTRATION
- Q10 value – reaction rate is doubled for - Small, non protein molecules (ex: vitamins)
every 10°C increase
- 40 – 50°C : enzyme undergoes inactivation J. PROSTHETIC GROUPS
and denaturation - Cofactors that bind tightly to proteins or enzymes
- Ex: Lipids, Carbs, metal ion
e. pH – Hydrogen Ion Concentration
- Changes in pH may denature an enzyme or
influence its ionic state resulting in structural
changes
- Optimum pH - the point at w/c the reaction
rate is greatest
- At pH 7.0 – 8.0, many enzymes show
maximum activity
- Pepsin: 1.5 and ALP at 10.5

f. ACTIVATORS: increase reaction rate

- Nonprotein entities that must bind to
particular enzyme
- -Bind the substrate to the active site by
forming ionic bridges
- Orients the substrate so it is attached to the
enzyme in the correct configuration
- Common activators are metallic (Ca, Fe, Mg,
Mn, Zn, K), inorganic, non-metallic (Br,Cl)

g. INHIBITORS
- Decrease the rate of enzyme reaction
- Interfere w/ the reaction

Competitive Inhibition - Inhibitors Binds to the active site,

blocks access of the S to the E (REVERSIBLE)

Non-competitive inhibition - Binds elsewhere on the E

causing change in shape that interferes w/ S binding (may be
reversible or irreversible(destroys) )

Uncompetitive inhibition - Inhibition-inhibitors binds to the ES

complex, if substrate will be increased, there will be increase in
ES conc. increasing inhibition, this inhibition does not yield
product.