HIV machinery structure cracked
Image: Salk Institute researchers, led by Dmitry Lyumkis (left), solve atomic structure of HIV machinery that has eluded researchers for decades.
Using cryo-electron microscopy, US-based Salk Institute researchers have solved the atomic structure of a key piece of machinery that allows HIV to integrate into DNA and replicate in the body.
The structure of this protein complex - known as 'intasome' - has eluded researchers for decades, but the latest information yields clues that will help researchers to develop new HIV drugs.
"We're particularly excited about the ability to understand and combat mechanisms of viral resistance," says Dr Dmitry Lyumkis from the Salk Institute. "HIV is a clever virus and has learned to evade even some of the best drugs on the market."
"Understanding the mechanisms of viral escape and developing more broadly applicable drugs will be a major direction in the future," he adds.
A key step in the life cycle of lentiviruses, such as HIV-1, is when viral DNA integrates into the host genome to infect the host cell.
This process is catalysed by the viral integrase enzyme, which is a major drug target.
As the virus integrates to the DNA, this enzyme binds the ends of viral DNA, forming the higher-order nucleoprotein complex, called the intasome.
Salk Institute scientists have solved the structure of the HIV intasome, a large molecular machine that inserts viral DNA into the genomes of its host. [Dario Passos and Jamie Simon/Salk Institute]
To solve the intasome structure associated with HIV-1, Lyumkis and colleagues turned to single-particle cryo-EM.
They first attached a specific protein to improve the intasome's ability to dissolve in liquid and then bathed the intasome in glycerol and salt to prevent particle aggregation.
Samples were applied to holey gold grids and then plunged into liquid ethane ready for cryo-EM analysis.
Researchers obtained cryo-EM data using a FEI Titan Krios electron microscope operating at 300 kV.
To overcome image contrast issues and ensure individual particles could be resolved, they used an exposure filter alongside a high electron dose, obtaining density maps resolved to ~3.5 to 4.5 Å.
Then, using a host of cryo-EM-related software, they produced 3D structures of the HIV intasome.
To the researchers' surprise, the HIV intasomes were more intricate and complex than other retroviruses.
Scientists already knew that the structures had a four-part core, but the latest results revealed that the HIV intasomes have many more units, known as "higher-order" species.
Evidence suggests that more complex versions of the intasome serve a purpose in helping HIV integrate itself within the host genome.
"HIV is like the luxury car whereas other retroviruses are the economy models--they're both cars, but the HIV intasome contains important upgrades to do different jobs," says Lyumkis.
The researchers' current study focused on intasomes after assembly on host DNA, but future work will study the structures prior to landing on the host genome and in the context of bound drugs.
To this end, the group is also working to push structure resolution ~4 to 2 Angstrom, providing crucial insight for drug discovery and development.
Research is published in Science.