Aspartate Proteases

RT Inhibitor	RT with Inhibitor Bound (1ep4)

The Inhibitor Does Not Bind in the Catalytic Site

Aspartate Proteases III

Two of the great plagues of humankind in the 20^th and 21^st centuries involve aspartate proteases as active participants: AIDS and Alzheimer's disease.

The AIDS virus has a single-stranded RNA genome, which is released after the virus enters the cell. The following outline is oversimplified, but correct in essence.

The RNA is then reverse transcribed into an RNA-DNA hybrid by the viral reverse transcriptase (RT) enzyme

Hence, the description of HIV as a retrovirus

The RT enzyme has no proofreading mechanism, and it is estimated that on average one mistake occurs for each reverse event

This results in an extremely high mutation rate for the HIV virus, and retroviruses in general

The RNA-DNA hybrid dissociates and the DNA can be converted to a double strand, and then possibly into the genome of the host organism; the RNA is reverse transcribed again

The viral genetic information codes chiefly for three large precursor proteins: gag, pol, and env
- env contains the envelope glycoproteins of the virus, and is processed normally by proteolytic enzymes of the host
- gag contains the proteins of the viral core and coat; pol encodes the reverse transcriptase, ribonuclease, and other enzymes of viral replication
- The remaining portion of the genome codes for the HIV protease, which cleaves the gag and pol proteins
This scheme suggests two methods for fighting the infection:
- inhibit the reverse transcriptase
- inhibit the protease
The first method tried was to inhibit the reverse transcriptase, which is a heterodimer, and has a catalytic site composed of three aspartates:

RT Inhibitor RT with Inhibitor Bound (1ep4)

The Inhibitor Does Not Bind in the Catalytic Site

It appears that HIV-RT inhibitors are allosteric inhibitors.
What is known about the mechanism of this enzyme is explicated in a recent review: J. Mol. Biol., 2009, 385, 693.
Our particular interest here is the protease, which is an extremely unusual aspartate protease.

HIV-1 Protease (side view), Catalytic Aspartates Shown HIV-1 Protease (top view)

What's unusual?
- The protease is composed of two identical chains
- The catalytic site is composed of one aspartate from each chain
- The chains are not covalently bound to each other
- The association is perfectly C₂ symmetric
No other protease has all four of these characteristics.
The tetrahedral intermediate has been caught in a flash-frozen crystal:

Note also the two large loops, called "flaps" at the top of the left picture.
- These are in the position shown, "closed" when an inhibitor is bound
- They are "open" when the catalytic site is empty
Structural Alignment of SIV, Open and Closed

The only "open" structure I could find is for one strand of the simian protease, so I aligned it with one strand edited from the pdb file of the complete virus bound to an inhibitor.
Although in this example it looks as if the flap has opened like a sliding roof, nmr evidence indicates that the flaps are fairly flexible.
This page last modified 1:16 PM on Thursday April 15th, 2010.
Webmaster, Department of Chemistry, University of Maine, Orono, ME 04469