U2 snRNP recognizes the branch site through a loaded-spring strand-invasion mechanism

Recognition of the branch sequence (BS) by the U2 snRNP is a pivotal step in pre-mRNA splicing and spliceosome assembly. Structural studies suggest that BS recognition occurs through a toehold-mediated strand-invasion mechanism, in which U2 snRNA progressively base-pairs with the intron to form the branch helix. However, given the limited complementarity between U2 snRNA and the intronic BS, it remains unclear how spontaneous strand invasion can occur. Here, using all-atom and coarse-grained molecular dynamics simulations, we show that strand invasion proceeds spontaneously once the toehold region is engaged and the TAT-SF1 factor is released. The key finding is that the branch-stem loop (BSL) of the U2 snRNA is maintained in a supercoiled, high-energy conformation by TAT-SF1, which acts as a molecular latch holding the BSL in a poised “loaded-spring” state. Displacement of TAT-SF1 allows the BSL to relax, releasing the stored conformational energy that drives strand invasion through local strand-slip and base-pair exchange. Moreover, the simulations reveal that strand invasion can proceed bidirectionally, refining previous models of U2–BS pairing. This work establishes a “loaded-spring” mechanism as a key physical driver of the toehold-mediated strand invasion underlying branchsite recognition within the early spliceosome.