College aantekeningen Biochemistry - Medical biochemistry and pathophysiology (5052MBP12Y)

Lecture 1 Biochemistry Enzymes

Michaelis Menten kinetic
1. Binding of substrate to substrate binding place on enzyme
a. Depends on affinity of enzyme and substrate  Km
2. Enzyme is catalyzing the reaction to modify the substrate
a. Is mostly irreversible
b. Catalysis  Kcat
c. Turnover number also depends also on V max

V = Vmax [S] / Kcat [S]

Enzyme contains about 500 amino acids. Only ~20 of them are used for the
binding side. And only ~3 would be used for actual catalysis.
If you mutualize the binding side the enzyme is nonfunctional since it can’t bind
any substrate

Only about 25% of human genome genes can encode enzymes

Inhibitors that binds reversibly to enzymes
- Competitive inhibitors
- Noncompetitive inhibitors

Inhibitors that bind irreversibly to enzymes
- Suicide inhibitors
o Mechanism based inhibitors
o Example is N,N-dimethylpropargulamine voor enzyme monoamine
oxidase (MOA). Normally it recognizes the drug as the substrate but
it is not a the substrate since it as a different group. And you get a
covalent bond. So it binds irreversibly and modifies the enzyme
thus becoming inactive. Is mechanism based since it depends on
the normal mechanism of the enzyme.

Allosteric enzymes and inhibitors
- Enzymes are multidomain enzymes or multisubunit enzymes such as
dimers
- Inhibitors bind allosteric enzymes reversibly al allosteric site and change
the enzyme conformation form a more-active, relaxed (R) state to less-
active tense (T) state

Cytochrome P450 enzymes (CYP)
- They all have the same mechanism, have different substrates to do it. Is a
protein that transfers electrons, using heme as prosthetic group
- Take a substrate and modify it into a OH group
o RH + O2 +NADPH + H  ROH + H2O + NADP
o Add two ions to make reaction happen. One added to substrate and
one used for H2O molecule

, - Enzyme = CYP
- Prosthetic group = heme
- Co-enzyme = NADPH
- Substrate = RH
- Co-substrate = O2




2

,Lecture 2 Biochemistry Drug development

Two approaches to drug discovery
1. Compound  physiological effect  molecular target
2. Molecular target  compound  physiological effect

Essential part of penicillin is the rectangle, which is the active group
Ampicillin has the same rectangle, only the side group is different.
Penicillin inhibits cross-linking of peptido-glycan chains, such as that the
bacterial cell wall weakens and bacterial cells lyse.
 The drug was found first and then the target was discovered

Sildenafil relaxes smooth muscle cells in blood veins which is controlled by cGMP.
So it is an enzyme inhibitor.

For treatment of AIDS multiple inhibitors are used. Since only one inhibitor is not
enough. The HIV virus will mutate and become resistant so multiple inhibitors
have a better effect. One possible inhibitor is the protease inhibitor which
prevents the chopping of the big protein into three workable proteins.
So they first looked at the substrate and then came up with a drug. Gave extra
groups to improve infinity of the compound so that the actual substrate won’t
bind.

Influenza H1N1 virus  H = hemaglutonin N = neurominidase
So H and N proteins on the surface that are necessary for binding, which comes
in many variations.
N cleaves the sialic acid residues from the glycoproteins at the cell surface to
release the viral particle, allowing it to infect another cell.
Made a drug that resembles the transition state of the sugars that binds to the
neuraminidase  binds so virus can’t bind anymore.

ADME properties of drugs  concentration of drug at target depends on it
- A = absorption  need to have balance of hydrophobic and hydrophilic so
it can cross the membrane
o Lipinski’s rules of 5
 Molecular weight < 500
 Number of H bonds donors < 5
 Number of H bond acceptors (N and O) < 10
 Partition coefficient, as log(P) < 5
 Log10 of the ratio of the drug concentration in octanol
to the concentration of water
- D = distribution
o Hydrophobic compounds do not dissolve freely in the blood but bind
to abundant proteins such as serum albumin
o Compounds distribute over various bode fluids and tissues, called
compartments
o Many compounds are unable to pass the blood brain barrier
- M = metabolism  little metabolism so it is not recognized by the liver




3

, o Defense of the body to modify compounds trough oxidation (phase
1) and conjugation (phase 2, really designate it to be removed)
o As result these compounds become more water soluble and more
easily recognized as foreign compounds
o Conjugation is addition of one of following groups to xenobiotic
compound
 Glutathione
 Glucuronic acid
 Sulfate
- E = Excretion  less as possible so it has change to work and so you don’t
have to take much of the drug
o Absorption through kidneys and excretion via urine
o Active transport by liver into bile and excretion into intestines and
stool
o Enterohepatic cycling  decreases rate of drug excretion because
some compounds are recycled after excretion
 From intestine to blood
 From blood to liver
 From liver via bile to intestine
o Drug has a half live (how long does it stay in the body)  t1/2


IC50 = inhibitory concentration at 50% inhibition
Log(P) = the partition coefficient
cmax = maximal concentration in the bloodstream
it is not necessarily the best inhibitor that is the best drug, also depends on the
cmax




4