The RAG recombinase drives V(D)J recombination and adaptive immunity, yet its evolutionary origin from a Transib-family transposon means it retains a latent transposase activity threatening genomic stability and contributing to chromosomal translocations and leukemias. Although molecular domestication has introduced suppressive adaptations, the mechanistic role of the RAG2 acidic hinge (AH), an intrinsically disordered region linking the Kelch and PHD domains remains poorly understood.
The AH was modeled in a fully extended state from the resolved RAG2 core using Modeller, then explored through 70 independent molecular dynamics simulations (100 ns each) with OpenMM, CHARMM36 force field, and implicit solvent at 310K. Trajectories were clustered to identify topological states and mapped onto the RAG tetramer surface.
Five recurrent conformational states provide a structural framework for understanding AH-mediated inhibition. In the most prevalent clusters, the AH localizes above the RAG1 DNA-binding groove, creating a steric and electrostatic barrier blocking target DNA acquisition and preventing the U-shape conformation required for transposition. A distinct cluster shows the AH intercalated at the lateral RAG1/RAG2 interface, acting as an allosteric wedge restricting inter-subunit flexibility essential for transposition. One particularly informative cluster shows the AH extended toward the RAG1 basic N-terminal region.
The acidic hinge thus emerges as a dynamic regulatory element whose electrostatic steering by basic surface patches positions it strategically to inhibit transposition and safeguard genomic stability during V(D)J recombination.

Co-authors: Eliza - Cristina Martin, Andrei - Jose Petrescu