Extended Data Fig. 9: RAD51, the eukaryotic RecA homologue, probably functions similarly to RecA in strand exchange.

RAD51 lacks the RecA CTD but instead contains an N-terminal domain (NTD) that has been implicated in dsDNA binding by chemical shift perturbation data of the isolated NTD domain³⁹. Although the RAD51 NTD is structurally unrelated to the RecA CTD, it occupies an analogous position at the filament periphery^40,41,42, except that it is oriented with its solvent-exposed surface pointing to the 5′ end of the filament instead of the 3′ end that the RecA CTD points to. Because of this, the NTDs at the 5′ end of the RAD51 filament are more accessible for initial dsDNA binding compared to those at the 3′ end, the opposite of the RecA filament. a, A side-by-side comparison of the RecA D-loop^DG structure (left) and a model of a 9-protomer RAD51–ssDNA presynaptic filament (right). The RAD51 model was constructed from the coordinates of the 4.4 Å cryo-EM structure of a 3-RAD51 segment bound to 9 nt of ssDNA (PDB: 5H1B) by successively applying the transformation that relates the middle RAD51 to the 5′-most RAD51. Because of small differences in the relative orientation of adjacent protomers, the RecA and RAD51 filaments are superimposable only locally (while individual helicase domains superimpose with a root-mean-square deviation (r.m.s.d.) of 1.9 Å for 201 of 245 RecA C_α atoms, three-protomer segments superimpose with an r.m.s.d. of 2.2 Å for 531 C_α atoms). For the side-by-side comparison, the two filaments were superimposed on the central, 3-protomer segment (RecA^D–RecA^F). The proteins and cofactors are coloured uniformly grey, except their respective CTD and NTD domains are coloured rainbow as in Fig. 1a, the primary ssDNA is brown for both, and the rest of the RecA DNA molecules are coloured as in Fig. 3a. The NTD and CTD domains are labelled. b, Magnified views of the RecA Dloop^DG and the 9-RAD51 model superimposed on the three 3′-most RecA^A–RecA^C, focusing on the RecA duplex^G in an orientation similar to a (right view rotated 90° about the vertical axis). This superposition brings duplex^G of RecA in close proximity to the RAD51 NTD^A, which is nearly one turn of the filament away (in the 3′ direction) from CTD^G owing to the different locations of the RAD51 NTDs. The backbone amide nitrogen atoms reported to be involved in dsDNA binding³⁹ are shown as yellow spheres (Ile61, Lys64, Gly65, Ile66 and Ala69). These are located at a loop (shown in thick tube) and the N terminus of the helix that follows. These structural elements are positioned relative to the DNA duplex analogously to the loop-helix motif of the RecA CTD, although they approach the DNA from the opposite direction. An adjacent RAD51 loop (residues 30–35, also shown as a thick tube) is also in close proximity to the RecA duplex^G of the superposition.