Dancing proteins: HIV targets

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  • Published: Oct 15, 2014
  • Author: David Bradley
  • Channels: X-ray Spectrometry
thumbnail image: Dancing proteins: HIV targets

Protein dance

New targets for preventative therapies of HIV infection might now be possible thanks to X-ray studies that reveal the 'dance' of HIV proteins on the surface of the virus that are thought to be involved in the infection of human immune cells. Image credit: Peter Kwong at the NIH

New targets for preventative therapies of HIV infection might now be possible thanks to new studies that reveal the 'dance' of HIV proteins on the surface of the virus that are thought to be involved in the infection of human immune cells. Single molecule imaging methods were used to reveal the dance of molecules. X-ray techniques were used to determine the structure of the closed conformation.

Parallel papers published in the journals Nature and Science reveal how US researchers at Weill Cornell Medical College have developed a technique that allows them, for the first time, to watch the behaviour of the human immunodeficiency virus type 1 (HIV-1) envelope (Env) spike, comprising three gp120 and three gp41 subunits. This protein is what the team describes as a "conformational machine". It eases entry of the virus into host cells - human CD4-carrying immune cells - by rearranging from a mature state with no ligand through receptor-bound intermediates, to a post-fusion state.

Understanding the details of this behaviour, or "dance", is down to the work of Scott Blanchard at Weill Cornell and his colleagues Walther Mothes, a HIV specialist at the Yale University School of Medicine, and James Munro, who was Blanchard's first graduate student and who is now an assistant professor at Tufts University School of Medicine and their respective teams. The Science paper discusses the dance while the Nature paper provides details of a three-dimensional X-ray structure of one of the conformations of the HIV protein.

Big questions, answered

Blanchard and his colleagues adapted an imaging technique that uses fluorescence to measure distance on molecular scale - single-molecule fluorescence resonance energy transfer (smFRET) imaging - to study the behaviour of the HIV spike protein. The development of novel fluorophores act as beacons when inserted into the outer protein coat, the envelope, of the virus; the fluorophores do not disturb the behaviour of the proteins. With two different fluorophores in place, the team used smFRET imaging to visualize the molecular dynamics as the viral proteins shift conformation. They demonstrated that the envelope, which consists of three gp120 and gp41 proteins positioned close together existing as a trimer, open up like a blooming flower in the presence of the immune cells' CD4 protein, exposing the gp41 subunit that is essential for subsequent aspects of the mechanism that leads to infection.

"Making the movements of HIV visible so that we can follow, in real time, how surface proteins on the virus behave will hopefully tell us what we need to know to prevent fusion with human cells - if you can prevent viral entry of HIV into immune cells, you have won," explains Blanchard. "What we have shown in the Science study is that we now have the means to obtain real-time images of processes happening on the surface of intact HIV particles, which we now plan to use to screen the impact of drugs and antibodies that can shut it down," he adds.

There are actually ten to twenty envelope trimers on the surface of each HIV virion. Moreover, these trimers mutate rapidly, which is partly the reason why HIV can so readily evaded a typical immune response and why it has proved impossible so far to develop a viable vaccine against the disease. The team explains that the gp120 proteins are constantly shape shifting, even when CD4 is not present. "This answered the first big question of how opening of the envelope trimer is triggered," Blanchard explains. "Many scientists believe that the particles remain in one conformation until they come across a CD4-positive cell. But we saw that the proteins dance when no CD4 was present - they change shape all the time." They are thus ready to infect the CD4 cells as soon as the opportunity arises. Indeed, the team introduced synthetic CD4 into the system they were studying and could then see how its presence stimulated the next step in what would normally be the infection process in a living person.

Viral intrigue

Intriguingly, the team also demonstrated that the presence of antibodies known to have some efficacy against HIV work by preventing gp120 from opening, which reduces the infection rate in the laboratory. In addition, a similar inhibitor effect occurs when an experimental small molecule drug now under development is introduced. "We are working now to improve the technology to achieve the imaging precision we need to make broadly effective therapies," Blanchard says.

Antibodies were also used to lock the protein conformation and stop the dancing in order to freeze the action for the X-ray crystallography work by Peter Kwong and colleagues reported in Nature. The protein constructs used in this investigation were originally developed by John Moore's team at Weill Cornell. "This concrete, atomic resolution picture of what the pre-fusion machinery looks like and where these antibodies bind provides an important step forward to understanding HIV's biology," Blanchard adds.

HIV has infected an estimated 70 million people around the world. And remains a pressing issue in medical research. The new research may well lead to a way to completely decode the infection processes for this and other retro viruses potentially facilitating the development of new pharmaceutical inhibitors.

Related Links

Nature 2014, online: "Structure and immune recognition of trimeric pre-fusion HIV-1 Env"

Science 2014, online: "Conformational dynamics of single HIV-1 envelope trimers on the surface of native virions"

Article by David Bradley

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

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