HIV Dynamics and Replication Program, National Cancer Institute, Frederick, MD 21702, USA.
Viruses. 2024 Sep 25;16(10):1517. doi: 10.3390/v16101517.
During HIV-1 virus assembly, the genomic RNA (vRNA) is selected for packaging by the viral protein Gag because it contains a specific packaging signal, Psi. While there have been numerous studies of Gag-Psi interactions, there is almost no information on the kinetic aspects of this interaction. We investigated the kinetics of Gag binding to different RNAs using switchSENSE DRX technology. We measured the association rate of Gag binding to monomeric Psi, to a "Multiple Binding Site Mutant" Psi that is inactive for genome packaging in vivo, and to a scrambled Psi. We discovered that Gag binds more rapidly to Psi RNA than to the mutant or scrambled RNAs. Furthermore, rapid Gag association kinetics are retained within sub-regions of Psi: Gag associates more rapidly with RNA containing only the 3' two of the three Psi stem-loops than with monomeric RNA containing the 5' two stem-loops or a scrambled RNA. No differences were detectable with individual Psi stem-loops. Interestingly, the rate of binding of Gag molecules to Psi increases with increasing Gag concentration, suggesting cooperativity in binding. The results are consistent with the hypothesis that selectivity in packaging derives from kinetic differences in initiation of particle assembly.
在 HIV-1 病毒组装过程中，基因组 RNA（vRNA）被病毒蛋白 Gag 选择进行包装，因为它包含一个特定的包装信号 Psi。虽然已经有许多关于 Gag-Psi 相互作用的研究，但几乎没有关于这种相互作用的动力学方面的信息。我们使用 switchSENSE DRX 技术研究了 Gag 与不同 RNA 的结合动力学。我们测量了 Gag 与单体 Psi、体内基因组包装无活性的“多结合位点突变体”Psi 以及乱序 Psi 的结合的缔合速率。我们发现 Gag 与 Psi RNA 的结合比与突变体或乱序 RNA 的结合更快。此外，Psi 的亚区域内保留了快速的 Gag 缔合动力学：Gag 与仅包含 3' 两个 Psi 茎环的 RNA 的结合比包含 5' 两个茎环的单体 RNA 或乱序 RNA 更快。单个 Psi 茎环没有可检测到的差异。有趣的是，Gag 分子与 Psi 的结合速率随 Gag 浓度的增加而增加，表明结合具有协同性。结果与这样的假设一致，即包装的选择性源于颗粒组装起始的动力学差异。
