Assembly of HIV-1, which causes AIDS, occurs on the inner plasma membrane leaflet of infected cells, an engineering construction process that creates hexagons of viral Gag protein trimers, as directed by Gag’s N-terminal matrix domain.
However, some details of virion assembly have been missing for four decades. In a study published in the journal Proceedings of the National Academy of SciencesJamil Saad and colleagues provided the first atomic demonstration of the matrix network, detailing molecular details at 2.1 Å resolution, a step that advances the understanding of the key mechanisms of viral assembly and viral envelope protein incorporation.
“Our findings may facilitate the development of new therapeutic agents that block HIV-1 aggregation, envelope incorporation, and ultimately virus production,” said Saad, professor of microbiology at the University of Alabama at Birmingham.
The Gag protein is modified after translation, in which a lipid-like myristate group is added to help Gag bind to the plasma membrane. How the myristoylated matrix domain, or myrMA, of Gag assembles into a network has so far eluded disclosure.
Technologies with low molecular precision -; such as cryo-electron diffraction and cryo-electron tomography; It was suggested that the myrMA protein is organized as trimers, and these trimers then undergo higher-order regulation to form hexagons of scissors. Saad’s study is consistent with a recent study, which indicated that the myrMA protein undergoes drastic structural changes to allow the formation of distinct hexagonal networks in immature and mature viral particles. Viral maturation is the final step in the viral replication cycle, in which a capsid core forms within the aggregated virus, yielding infectious particles.
HIV-1 envelope protein, or Env, is a transmembrane protein that is delivered to the plasma membrane by the cell’s secretory pathway. The bulk of the Env protein extends beyond the membrane, but the tail hangs across the membrane back into the cell. Genetic and biochemical studies suggested that incorporation of the viral Env protein into virus particles also depends on the interaction between the myrMA domain and the cytoplasmic tail of Env. In 2017, Saad’s lab resolved the high-resolution structure of the cytoplasmic tail of Env, which was the last unknown protein structure of HIV-1.
Env is a key infection protein. When mature HIV-1 approaches the target cell, Env attaches to proteins on the outside of the uninfected cell, and the Env protein then bursts like a mousetrap to fuse the viral membrane with the cell membrane.
In the structures described by Saad and UAB colleagues, myristic myrMA plays a major role in stabilizing the reticular structure, so the ability to form myrMA crystals was important. They achieved this elusive technical challenge by removing 20 amino acids from the end of myrMA’s 132 amino acids. It is known that the formation of a Gag network on the plasma membrane is mandatory for assembly of immature HIV-1 and incorporation of Env.
Saad and colleagues report that their myrMA network is arranged in a hexagonal form of trimmers with a central hole, believed to accommodate the C-terminal tail of Env to promote integration into viruses. Their myrMA crystals allowed them to monitor the myr group connected in the network. They find that the myr group of one myrMA subunit is inserted into the hydrophobic cavity of the subunit across the two-slit axis, presenting a ‘myristoyl swap’, and they also report other molecular interactions between trimers. The researchers describe additional molecular details that help stabilize the hexagonal shape of the shear network.
By performing paired mutation studies with nuclear magnetic resonance, or NMR, researchers have provided evidence that a single amino acid substitution in the matrix -; Leucine-13 or Leucine-31 for glutamic acid -; caused a conformational change in myrMA that might destabilize the trimmer interactions within the network. Previous genetic studies indicated that substituting Leucine-13 or Leucine-31 has deleterious effects on Env incorporation.
Another important finding in this study is the evidence for the alternating membrane-binding mechanism of Gag, which is known to be mediated through interactions of the myrMA domain with phosphatidylinositol 4,5-bisphosphate, or PI(4,5)P.2, Topical lubricant exclusively on the inner leaflet of the plasma membrane. UAB researchers showed that PI(4,5)P.2 Able to link to alternative sites on the MA. This corresponds to a novel mechanism of alternating MA-membrane binding PI(4,5)P .2 During aggregation of immature particles and during maturation.
“In conclusion, we provided an atomic view of the HIV-1 myrMA network that revealed invaluable structural insights into the arrangement of myrMA subunits, trimers, the trimer-trimer interface, the Myr swap, and the effect of defective MA mutations in the Env fusion on the structure,” Saad said. mirma plant and thus reticular formation.” “Our data also supported the alternating MA–PI(4,5)P2 Mechanism of splicing during virus assembly and maturation. These results filled a significant gap in our understanding of the mechanisms of Gag assembly on the plasma membrane and Env incorporation into virus particles.”