Qβ類病毒顆粒蛋白包裹23S核糖體核糖核酸之研究

Studies of the 23S rRNA packed in the Qβ virus-like particles

張耘誌(Yun-Chih Chang)

指導教授 ： 蕭超隆

國立臺灣大學/生命科學院/生化科學研究所/碩士(2021年)

https://doi.org/10.6342/NTU202100036

摘要

噬菌體Qβ 的外殼蛋白二聚體能藉由與「operator」核醣核酸的交互作用，進而形成類病毒顆粒蛋白。利用這項特性，可將欲包裹的分子透過「operator」 核醣核酸置入類病毒顆粒蛋白。這邊，我們測試Qβ 類病毒顆粒蛋白能否包覆23S 核醣體核糖核酸分子。我們假設23S 核醣體核糖核酸分子只有在「operator」核糖核酸分子的存在下能被Qβ 類病毒顆粒蛋白包裹。為測試假設，我們製備了兩種型態的23S 核醣體核醣核酸：（1）「operator」核酸不存在的23S 核醣體核醣核酸，（2）「operator」核醣核酸建構於23S 三端的核醣體核醣核酸。實驗結果顯示上述的兩種23S 核醣體核醣核酸皆能被Qβ 類病毒顆粒蛋白包裹。根據先前的研究，「operator」核醣核酸的變異體在結構與序列上有三樣特性（1）核酸主幹（stem）至少有六組鹼基對、（2）環圈（loop）部分需有三或四個核酸苷、（3）環圈部位的最後一個鹼基必須是腺嘌呤。依上述條件，我們在23S 核醣體核糖核酸的一百零一個螺旋（helix）中發現有八個螺旋滿足其特徵。這八個狀似「operator」核醣核酸的核醣體螺旋裡，有五個螺旋的環圈是位向溶劑可接觸面。依此推測，可能是促成Qβ 類病毒顆粒蛋白組裝的關鍵螺旋。因此，23S 核醣體核醣核酸是可以在「operator」核醣核酸不存在的條件下完成Qβ 類病毒顆粒蛋白的組裝。

關鍵字

Qβ 類病毒顆粒蛋白 ； 23S 核醣體核糖核酸 ； 「operator」核醣核酸

並列摘要

The bacteriophage Qβ coat protein dimers form virus-like particles (VLPs) through “operator” RNA interactions. With this feature, the molecular of interest can be packed into VLPs. Here, we test whether the Qβ VLPs can encapsulate the 23S rRNAs. We hypothesized that the 23S rRNAs could only be packed into the Qβ VLPs in the presence of the “operator” RNA. To test our hypothesis, we prepared two types of the 23S rRNA: (i) the 23S rRNA contained no “operator” RNA, and (ii) the 23S rRNA with the “operator” RNA constructed at the 3’-terminus. The results show that both the 23S rRNAs can be packed in the Qβ VLPs. Previous studies have shown that the structure and sequence of the “operator” RNA variants are featured: (i) at least six base pairs presented on a helical stem, (ii) three or four nucleotides composed a terminal loop, and (iii) the last residue required a nucleobase A in the terminal loop region. Accordingly, of the 101 helices within the 23S rRNA, eight helices exhibit the “operator” RNA’s characteristics. Furthermore, among those eight “operator”-like rRNA helices, the five helices of which their loop regions are shown solvent-accessible in 3D, suggested that these five helices may be critical for the assembly of the Qβ VLPs. We concluded that the Qβ VLPs encapsulate the 23S rRNA independent of the “operator” RNA.

並列關鍵字

Qβ virus-like particle ； 23S rRNA ； “operator” RNA

參考文獻

1. Rumnieks J, Tars K: Crystal structure of the bacteriophage Qbeta coat protein in complex with the RNA operator of the replicase gene. J Mol Biol 2014, 426(5):1039-1049.

2. Weber H: The binding site for coat protein on bacteriophage Qβ RNA. Biochimica et Biophysica Acta (BBA)-Nucleic Acids and Protein Synthesis 1976, 418(2):175-183.

3. Witherell GW: Specific Interaction between RNA Phage Coat. Progress in nucleic acid research and molecular biology 1991:185.

4. Roldao A, Mellado MCM, Castilho LR, Carrondo MJ, Alves PM: Virus-like particles in vaccine development. Expert review of vaccines 2010, 9(10):1149-1176.

5. Witherell GW, Uhlenbeck OC: Specific RNA binding by Q beta coat protein. Biochemistry 1989, 28(1):71-76.