Structure and Mechanical Stabilities of the Three-Way Junction Motifs in Prohead RNA.

The core structure of phi29 prohead RNA (pRNA) is composed of three major helices organized into three-way junction pRNA (3WJ-pRNA) and has stout structural rigidity along the coaxial helices. Prohead RNAs of the other Bacillus subtilis bacteriophages such as GA1 and SF5 share similar secondary structure and function with phi29; whether these pRNAs have similar mechanical rigidity remains to be elucidated. In this study, we constructed the tertiary structures of GA1 and SF5 3WJ-pRNAs by comparative modeling. Both GA1 and SF5 3WJ-pRNAs adopt a similar structure, in which three helices are organized as the three-way junction and two of the three helices are stacked coaxially. Moreover, detailed structural features of GA1 and SF5 3WJ-pRNAs are also similar to those of phi29 3WJ-pRNA: all of the bases of the coaxial helices are paired, and all of the adenines in the junction region are paired, which eliminates the interference of A-minor tertiary interactions. Hence, the coaxial helices tightly join to each other, and the major groove between them is very narrow. Two Mg2+ ions can thus fit into this major groove and form double Mg clamps. A steered molecular dynamics simulation was used to study the mechanical properties of these 3WJ-pRNAs. Both GA1 and SF5 3WJ-pRNAs show strong resistance to applied force in the direction of their coaxial helices. Such mechanical stability can be attributed to the Mg clamps.