Mechanism of three-component collision to produce ultrastable pRNA three-way junction of Phi29 DNA-packaging motor by kinetic assessment
Mechanism of three-component collision to produce ultrastable pRNA three-way junction of Phi29 DNA-packaging motor by kinetic assessment
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DOI：
10.1261/rna.057646.116
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发表时间：
2016-11-01
期刊：
影响因子：
4.5
通讯作者：
Guo, Peixuan
中科院分区：
文献类型：
--
作者：
Binzel, Daniel W.;Khisamutdinov, Emil;Guo, Peixuan
RNA纳米技术正在迅速兴起。由于有利的药代动力学和有利的体内生物分布，RNA纳米颗粒在靶向递送治疗剂中显示出前景。RNA纳米技术应用自下而上的组装，因此阐明多个组分之间的相互作用机制是至关重要的。随着phi 29 DNA包装马达的新型热稳定性三向连接（3 WJ）基序的发现，人们对RNA不稳定性的关注逐渐减少。研究了这三种组分的动力学，每种组分平均18个核苷酸（nt），以阐明产生稳定的3 WJ的机制。三个片段通过两步反应机制以惊人的速度和亲和力组装成3 WJ：3 WJ（B）+3 WJ（c）3 WJ（bc）+3 WJ（a）3 WJ（abc）。3 WJ（B）和3 WJ（c）之间反应的第一步是高度动态的，因为这两个片段仅含有8个核苷酸用于互补。在第二步中，含有与3 WJ（bc）复合物互补的17个核苷酸的3 WJ（a）将不稳定的3 WJ（bc）复合物锁定为高度稳定的3 WJ。所得pRNA-3 WJ比任何二聚体种类更稳定，如更快的缔合速率和最慢的解离速率常数所示。第二步发生在一个非常高的缔合速率，这是难以量化的，导致快速形成稳定的3 WJ。阐明了三组分碰撞产生超稳定3 WJ的机制，证明了RNA纳米颗粒自下而上组装作为一类新的阴离子聚合物用于材料科学，电子元件或治疗试剂的有前途的平台。
RNA nanotechnology is rapidly emerging. Due to advantageous pharmacokinetics and favorable in vivo biodistribution, RNA nanoparticles have shown promise in targeted delivery of therapeutics. RNA nanotechnology applies bottom-up assembly, thus elucidation of the mechanism of interaction between multiple components is of fundamental importance. The tendency of diminishing concern about RNA instability has accelerated by the finding of the novel thermostable three-way junction (3WJ) motif of the phi29 DNA-packaging motor. The kinetics of these three components, each averaging 18 nucleotides (nt), was investigated to elucidate the mechanism for producing the stable 3WJ. The three fragments coassembled into the 3WJ with extraordinary speed and affinity via a two-step reaction mechanism, 3WJ(b) + 3WJ(c) 3WJ(bc) + 3WJ(a) 3WJ(abc). The first step of reaction between 3WJ(b) and 3WJ(c) is highly dynamic since these two fragments only contain 8 nt for complementation. In the second step, the 3WJ(a), which contains 17 nt complementary to the 3WJ(bc) complex, locks the unstable 3WJ(bc) complex into a highly stable 3WJ. The resulting pRNA-3WJ is more stable than any of the dimer species as shown in the much more rapid association rates and slowest dissociation rate constant. The second step occurs at a very high association rate that is difficult to quantify, resulting in a rapid formation of a stable 3WJ. Elucidation of the mechanism of three-component collision in producing the ultrastable 3WJ proves a promising platform for bottom-up assembly of RNA nanoparticles as a new class of anion polymers for material science, electronic elements, or therapeutic reagents.