REGULATION OF ENZYME ACTIVITY iii.

Urea cycle enzymes

A. PATHWAY CONTROL – modulation of activity
of one or more key enzymes in a pathway 2. Degradation
1. Rate-limiting enzyme – the enzyme a. Ubiquitin-Proteasome pathway –
with the lowest vmax in the pathway; degradation of proteins tagged by
enzymes that catalyze the “bottleneck” or ubiquitin in the proteasome
rate-limiting reaction ● 26S proteasome – large
- decrease in catalytic efficiency or macromolecular complex in the
quantity of the catalyst = shape of a hollow cylinder; active
reduction in metabolite flux or sites in the interior part
decrease in rate (slow down) ● Ubiquitin – small (~8.5 kDa)
- increased quantity or catalytic protein; highly conserved in
efficiency = increase in flux eukaryotes
through the pathway (ex: HMG ● Ubiquitination – process of
CoA reductase – rate-limiting tagging misfolded and
enzyme in cholesterogenesis or functionally-compromised
“statin” drugs inhibit cholesterol proteins (including enzymes)
synthesis since it inhibits HMG catalyzed by E3 ligases that
CoA reductase) attach ubiquitin to the side-chain
2. Enzyme associated with the amino group of lysyl residues of
committed step: enzyme that catalayze the target
the 1st irreversible reaction in the - Covalent modifications
pathway - Binding with allosteric effector/substrate
- Short-term regulation: regulation happens via - Association with membrane
activity modification of the existing enzyme oligonucleotides or other proteins
without change in concentration
- Long-term regulation: refers to changes in Control of Enzyme Catalytic Efficiency
amount of enzyme without altering its kinetic - directly controlled through
properties conformational or structural alterations
- Cross-regulation: the product of pathway1 is an - Can be modulated by activators,
inhibitor or activator of an enzyme in pathway2 inhibitors or via covalent modifications
A. Allosteric Regulation
B. MORE REGULATION OF ENZYME 1. Feedback inhibition via negative
ACTIVITY allosteric effector
Control of Enzyme Concentration ● competitive,
- Control of enzyme availability: will depend on noncompetitive or mixed
the rate of synthesis and rate of degradation ● Typically inhibits the 1st
- Can be regulated by the changes in de novo committed step in the
synthesis of the enzyme biosynthetic pathway
1. Synthesis 2. Multiple feedback loops
a. Repressors: via excess (Cooperative Feedback
metabolites Inhibition)
b. Stimulators: via hormones and ● Effect may be strictly
other EC signals additive
c. Inducers (for inducible enzymes) ● Effect may be greater
i. Cytochrome P450 than the individual
(detoxification) feedback loop
ii. HMG-COA reductase 3. Release of Allosteric 2nd messenger
(cholesterol synthesis)
 ● 1st messenger: hormones or nerve impractical energy-wise
impulse o restoration to original state occurs via a different
● 2nd messenger: cyclic AMP reaction
B. Covalent Modification o e.g. phosphorylation transfer of Pi
1. Irreversible – Partial proteolysis to protein, catalyzed by protein kinase;
● For enzyme activation dephosphorylation hydrolytic reaction, catalyzed
● Enzymes for blood clotting protein phosphatases
(thrombin) and powerful
hydrolytic digestive enzymes are Compartmentalization
secreted in the proenzyme or Physically partitioning the pathway from its initial
zymogen form substrate by controlling the access of substrate to
● Example: removal of amino the enzymes of a pathway
terminal of pepsinogen results to A. Specific subcellular compartment
pepsin ● Lysosomes: enzymes degrade proteins
o For protection from and polysaccharides
autodigestion ● Cytosol: enzymes for FA synthesis
o Facilitates rapid ● Mitochondria: FA oxidation
mobilization in response B. Specialized cell type compartment
to demand C.Thermodynamic substitution (ex. Kinase-
2.Reversible – phosphorylation phosphatase): substitution of 1 or more reaction
dephosphorylation: most common by different reaction favored thermodynamically
● permits the functional properties in the opposite direction
of the affected enzyme to be D.Discrimination(NAD:catabolism;
altered only for as long as it NADP:anabolism)
serves a specific need
● the high charge density of the Multimeric Ezymes
phosphoryl group and their - Highly organized systems
propensity to form strong salt - A state of aggregation involving several
bridges renders them potent different enzymes (Example: pyruvate
agents for modifying protein dehydrogenase complex and α-
structure and function ketoglutarate dehydrogenase complex)
● extremely versatile because it - Product of first enzyme becomes
alters: substrate of the next enzyme in the
i. enzyme location in cell series
ii. enzyme susceptibility to - Each component is arranged to afford
degradation efficient coupling of individual reactions
iii. enzyme catalytic in-/efficiency catalyzed by the enzymes
iv. enzyme responsiveness to - Increases reaction rates and eliminates
allosteric regulation side reactions = increased overall
● other forms: acetylation, efficiency
methylation, ADP-ribosylation
● reversible because the modified Presence of Isozymes
protein can be restore back to its - Physically distinct versions of a given
original, modification-free state enzyme (catalyze the same reaction)
o does not mean - Products of gene duplication (altered at
reversing the reaction certain amino acids)
o if modification = - Exhibit subtle differences in properties
thermodynamically favorable, - Confer tissue specificity and adapt the
then simply reversing the process = unfavorable, tissues to special circumstances
 - Provide a “back-up” mechanism iii. LDH 3 (H 2 M 2 ) – brain and
- Example: kidney tissue injury
- Glucokinase (liver) vs iv. LDH 4 (HM 3 ) – skeletal
hexokinase (brain & muscle injury
muscle) ● v. LDH 5 (M 4 ) – liver cell injury
● Allows the tissue to (acute hepatitis)
adapt to special
circumstance
● Affect the reaction:
glucose to glucose-6-
phosphate
● Hexokinase- active all
the time (steady supply
of glucose for the brain)
● Glucokinase- active only
after meal (high blood
sugar)
- Creatine Kinase
● Allows tissue specificity Figure 1. Extent of Enzyme Increase (Dr. Menorca’s ppt)
(MM-muscle, BB-brain,
MB- heart)
2. Reagents
- Enzymes can be used for screening test
CLINICAL APPLICATIONS for cholesterol and triglycerides for a few
1. Diagnostics minutes using 10mL of plasma
- Plasma enzymes - Cholesterol oxidase and lipase are the
● High concentration in the active components of the assay system
plasma - The enzymes are immobilized in a bilayer
● Specific and has a functional along the necessary buffer
role in the plasma salts,cofactors or cosubstrates and
- Non-functional plasma enzymes indicator agents
● Present in plasma at low levels - Glucose oxidase (for blood sugar) can be
● No functional role in the plasma embedded on a strip of paper. The strip
- Measurement of plasma enzyme activity of paper will change its color through the
● Non-plasma enzymes: specific action of the embedded enzyme. Blood
enzymes sugar level can be determined by
- Increase in non-plasma enzymes comparing the color on the strip of paper
indicate that there is a diseased or to a reference standard. This can be
problematic tissue or organ done in just a minute.
● CREATINE PHOSPHOKINASE - This is important for those who monitor
i. MB CPK – Acute M.I the blood sugar of patients with diabetes
ii. MM CPK – Skeletal Muscle especially those with hyperosmolar coma
iii. BB CPK – Brain Tissue or diabetes ketoacidosis.
● ALKALINE PHOSPHATASE - Monitoring should be done hourly. With
(Obstructive jaundice (CBD this, you can reverse the situation as fast
stone; Cirrhosis; Cancer) as you could
● LACTATE DEHYDROGENASE
i. LDH 1 (H 4 ) – acute M.I.
ii. LDH 2 (H 3 M) – acute M.I.
 References
Dr. Menorca’s PPT
Previous transes (2022 A, B, C, D)
Rodwell, V., Kennelly, P., Bender, D., Weil, P.A.,
& Botham, K. (2018). Harper’s Illustrated
Biochemistry (31st ed).
Fig 2. Enzymes (bold) used as reagents (Dr. Menorca’s ppt)

- ENZYME-LINKED IMMUNOSORBENT
ASSAY (ELISA)
● A plate-based assay technique
designed for detecting and
quantifying substances such as
peptides, proteins, antibodies
and hormones
● Enzymes makes the
determination more specific.
● Combine enzyme action with
antigen-antibody interaction.

Figure 3. ELISA (Dr. Menorca’s PPT)

3. Therapeutic Agents
- Streptokinase (Useful in clearing blood
clot that occur in M.I. and lower E.)
- Plasmin (A serine protease that cleaves
the insoluble fibrin in blood clots into
several soluble components)
- Asparaginase (Used in the treatment of
some adult-type leukemia
- Enzyme replacement therapy ( In the
future, medical treatment replacing an
enzyme in patients in whom that
particular enzyme is deficient or absent)