HIV1 Protease and "Flap" Orientations

Enzymes are the machines of the cell - they make almost all of the chemical reactions that take place within a cell happen in a realistic time scale. They are able to do this because they bind specifically to a target molecule (the substrate) and convert it into a new molecule.

There are two models that are frequently used to describe how this "binding" works. The simplest is the Lock-and-Key model which assumes that the enzyme is a rigid molecule with a hole in it, rather like a lock and its keyhole. If a key has the right size and shape to fit into the keyhole, it might be able to open the lock. The enzyme has an opening called the active site - molecules with the right size and shape can fit into this active site and be modified by the enzyme. A drug (an inhibitor) can be designed that has the right size and shape to fit into the active site, but then it gets stuck. Once the active site is blocked by the inhibitor, the enzyme can no longer convert substrate molecules and it no longer works.

This model is rather limited - enzymes are often quite flexible. The second model, called induced fit, says that the enzyme changes shape when it binds to the substrate or inhibitor. If this happens, it will be important to know not only what the active site is like, but you also need to know how the enzyme changes when it binds to the substrate or inhibitor if you want  to design an effective drug.

Two recent papers examine how binding to an inhibitor may affect the shape of HIV-1 protease. HIV-1 protease (HIV PR) is an essential enzyme in the functioning of HIV and the target of many drugs for treating AIDS. If HIV-1 protease can be inhibited, none of the other proteins needed by HIV will get processed into their active forms.

HIV-1 protease bound to an inhibitor. Image from http://en.wikipedia.org/wiki/File:Hiv-1_pdb_1ebz.png

Drug Pressure Selected Mutations in HIV-1 Protease Alter Flap Conformations looks at mutations in HIV-1 protease.  When HIV is exposed to protease inhibitor drug cocktails they observe mutations in HIV PR, especially in the two loops or flaps that cover the active site.  Mutations that reduce the ability of the drug to bind to the enzyme active site will be resistant to the influence of the drug.  Changes in the active site itself would obviously have an affect on the ability of the drug to bind effectively, but why would the protein develop mutations in the flap region, which is not directly related to the active site?  By looking at the orientation of the flaps when different inhibitors are bound to the mutants, they suggest that the flaps adjust to accomodate the binding of the substrate/inhibitor in order to fine tune the binding strength.

The second paper, Dynamics of “Flap” Structures in Three HIV-1 Protease/Inhibitor Complexes Probed by Total Chemical Synthesis and Pulse-EPR Spectroscopy, also looks at the flaps and inhibitor binding.  They also see evidence that the flaps move in response to the nature of the substrate as it is bound to the enzyme.  The reaction catalyzed by HIV PR involves two distinct chemical steps, so they chose inhibitors that resembled different stages along the reaction sequence:  during the first step, between steps one and two, and during step two.  They conclude that the flaps move to fine-tune the interaction between the enzyme and the substrate as the reaction procedes.

Both papers report evidence of "induced-fit" behavior in the way HIV PR interects with it's substrate.  Understanding what role the flaps play in substrate binding can lead to better drug s for treating Aids.

Luis Galiano, Fangyu Ding, Angelo M. Veloro, Mandy E. Blackburn, Carlos Simmerling, Gail E. Fanucci (2009). Drug Pressure Selected Mutations in HIV-1 Protease Alter Flap Conformations Journal of the American Chemical Society, 131 (2), 430-431 DOI: 10.1021/ja807531v

Vladimir Yu. Torbeev, H. Raghuraman, Kalyaneswar Mandal, Sanjib Senapati, Eduardo Perozo, Stephen B. H. Kent (2009). Dynamics of “Flap” Structures in Three HIV-1 Protease/Inhibitor Complexes Probed by Total Chemical Synthesis and Pulse-EPR Spectroscopy Journal of the American Chemical Society, 131 (3), 884-885 DOI: 10.1021/ja806526z