OC 231 Assignment 2

Q. Get the structure of HIV protease dimer. Write about the peptide hydrolysis mechanism of HIV
protease 1.
Ans. HIV, like many other viruses, makes proteins in one long stretch, and it is the job of the HIV-1
protease to chop these into individual proteins, so that they can form a mature virus and infect
new cells.
The protein consists of 2 identical protein chains that are 99 amino acids long. It is an aspartyl
protease. Each monomer consists of an extended -sheet region, which is glycine rich, known
as the flap. This contributes to the substrate-binding site. One of the 2 essential aspartyl
residues, Asp 25 and Asp 25 lie on the bottom. This protease has the sequence Asp-Thr-Gly,
which is conserved in many aspartyl proteases.
The protein uses an activated water molecule to attack the amide bond carbonyl of the Phe-Pro
peptide bond. The activation of the water molecule is achieved by the aspartyl -carboxy
groups at the active site, which is characteristic of Aspartyl proteases.
Jasklski et al have proposed a mechanism based on the crystal structure a chemically synthesized
HIV-1 protease and an octapeptide inhibitor. The hydrolysis reaction proposed is a one-step
process during which the nucleophilic water molecule and the acidic proton attack the PhePro peptide bond in a concerted manner.

This is the proposed concerted mechanism for peptide hydrolysis by HIV-1 protease.

Another mechanism was proposed by Hyland et al based on pH-rate, 18O incorporation and
other methods. In this mechanism, Asp-25 exists in the unprotonated state upon binding to the
substrate, while the Asp-25 proton is H-bonded to the carbonyl oxygen of the substrate. The water
molecule is positioned closer to the -carboxy group of Asp-25. It has some resemblance to the
concerted enzyme, in that the amine product is protonated by an active site Asp. In this mechanism, a
discrete tetrahedral intermediate is formed, which was lacking in the concerted mechanism.
However, there are still many questions that are left to be answered about the catalytic
mechanism of HIV-1 protease. But these studies have led to the development of HIV protease
inhibitors, 6 of which have been FDA approved.

This figure shows the mechanism proposed by Hyland et al.

This figure shows the structure of HIV-1 protease, complexed to the inhibitor TL-3.
References: Brik, A. and Wong, C.H., 2003. HIV-1 protease: mechanism and drug discovery. Organic
& biomolecular chemistry, 1(1), pp.5-14.

Q. Draw structures of 10 peptide bond isosteres.

Ans. Ester, thioester, alkene, fluoroalkene, silanediol, aminoethylene, trifluoroethylamine,
ketomethylene, hydroxyethylene, dihydroxyethylene, hydroxyethylamine, methyleneamine etc. 10 of
these are shown below.

Figure 2: An ester
Figure 1: A peptide bond

Figure 3: A thioester

Figure 5: A fluoroalkene
Figure 4: An (E)-alkene (non-hydrolyzable isostere)

Figure 6: A silanediol

Figure 7: An aminoethylene group

Figure 8: A trifluoroethylamine group

hydrolyzable)

Figure 9: A ketomethylene group (non-

Figure 10: A hydroxyethylene group (non-hydrolyzable)

Figure 11: A hydroxyethylamine group (non-hydrolyzable)

References: Choudhary, A. and Raines, R.T., 2011. An Evaluation of PeptideBond

Isosteres. ChemBioChem, 12(12), pp.1801-1807.
Wipf, P., Xiao, J. and Stephenson, C.R., 2009. Peptide-like molecules (PLMs): a journey from peptide
bond isosteres to Gramicidin S mimetics and mitochondrial targeting agents. CHIMIA International
Journal for Chemistry, 63(11), pp.764-775.