Rational drug design

Question 1

What are the surface proteins (spikes) of influenza A ?

Answer 1

• Hemagglutinin (HA)
• Neuraminidase (NA)

Question 2

Name the five steps of the infectious cycle of a virus.

Answer 2

1. Attachment of virions to host cells
2. Endocytosis
3. Replication, translation, posttranslational processing
4. Budding (Keimung)
5. Release

Question 3

What is the function of Hemagglutinin (HA) at the surface of influenza A ?

Answer 3

• HA bind to sialic acid residues
• Attachment to host cell surface

Question 4

What is the function of Neuraminidase (NA) at the surface of influenza A ?

Answer 4

• NA cleaves sialic acid residues
• Release of new viruses from the host cell surface

Question 5

Which sialic acid can often be found at the end of carbohydrates ?

Answer 5

Neu5Ac

Question 6

Which bond can usually be found between Neu5Ac and other monosaccharides ?

Answer 6

O-glycosidic bond
- α-2,3 or
- α-2,6

Question 7

Which kind of structure could be used to block hemagglutinin (HA) ? Explain your answer.

Answer 7

• HA binds Neu5Ac at the cell surface with a high affinity
• A structure similar to Neu5Ac could be used for blocking HA

Question 8

Which kind of structure could be used to block neuraminidase (NA) ? Explain your answer.

Answer 8

• NA has a low affinity to sialic acids and cleaves its glycosidic bond
• A structure similar to the transition state of the enzymatic cleavage reaction could be used for blocking NA

Question 9

In 1970 a neuraminidase inhibitor was developed. Which one was is and how high was its affinity in contrast to Neu5Ac ?

Answer 9

Inhibitor: Neu5Ac2en
- Similar structure to the normal substrate Neu5Ac
- Affinity of Neu5Ac: K_i = 10^-3M
- Affinity of Neu5Ac2en: K_i = 10^-6M
- Affinity of inhibitor is 100-fold higher

Question 10

In 1993 a second neuraminidase inhibitor was developed. Which one was it, how was it different from Neu5Ac2en and how high was its affinity to neuraminidase ?

Answer 10

Inhibitor: Zanamavir
- In contrast to Neu5Ac2en, Zanamavir has a basic residue in the fourth position
- Affinity of Zanamavir: K_i = 10^-10M to 10^-11M

Question 11

What is log P ?

Answer 11

• A scale that indicates the solubility of a molecule
• P is the partition constant
• P is defined as the ratio of concentration of a compound in aqueous phase to the concentration in an immiscible solvent

Question 12

How is log P calculated ?

Answer 12

log P = log_10 (OP/AP)

Question 13

Tamiflu is an improved neuraminidase inhibitor. Name its modifications and their advantages.

Answer 13

• 3-Pentyl ether improves lipophilicity and complementarity to binding pocket
• Remoded oxygen leads to better stability
• Corrected position of the double bound mimic the transition state more successful
• Ester increases the lipophilicity
• Basic residue interacts with Glu 119 and Glu227

Question 14

How is the dissociation constant K_D calculated ?

Answer 14

K_D = (c(P)*c(L)) / c(PL) = koff/kon

Question 15

How is the dissociation constant K_D determined graphically ?

Answer 15

• The reaction response is plotted against the substrate concentration
• The substrate concentration at 1/2 of the maximum reaction response is K_D

Question 16

What is K_i ?

Answer 16

• Inhibition constant
• Says something about the affinity of an inhibitor -> The smaller K_i, the higher the affinity
• K_i is the inhibitor concentration at 1/2 of the maximum inhibition of the reaction
• K_i is often in the same order of magnitude as K_D

Question 17

Which value has K_i of Tamiflu for the inhibition of influenza A ?

Answer 17

~ 10^-11 M

Question 18

Explain one method to synthesize a ester library.

Answer 18

Parallel synthesis
- A few different alcohols and a few different acid chlorides are used
- They are made to react in each combination to form esters
- Method can be applied with huge numbers of different compounds by robots under computer control

Question 19

What is PEA ?

Answer 19

• Proximity Extension Assay
• Fast screening method for specific proteins
• Screen for quality but only approx. quantity of proteins

Question 20

How does a proximity extension assay work ?

Answer 20

• Antibody pairs labelled with a complementary sequence of DNA oligonucleotides are synthesized
• Antibodies bind pairwise to the analytes
• DNA oligonucleotides hybridize and extend
• Screening for specific analytes using quantitative polymerase chain reaction (qPCR)

Question 20

What happens to the equilibrium of an enzymatic reaction if the enzyme binds the reaction product ß

Answer 20

• Reaction product is removed from the ongoing reaction
• Equilibrium shifts towards the product

Question 20

How is the Gibb´s free energy ΔG calculated ?

Answer 20

ΔG = -RT*ln(K)