Figure 2

Idealized schematic representations of synaptic complexes (SC I and SC II) of circle-forming insertion sequences. Each complex is shown as a dimer (aqua ovals) with a BS (orange) and a CC (purple). Each IR is complexed with its PBD (red for IRR and blue for IRL) to the BS of its monomer, and its CD bound in cis to the CC. (A) In SC I, at one stochastically activated CC (IRR in this case) the CD is cleaved at its 3' end, exposing a 3'OH group (black half arrow) which, in a transesterification reaction, attacks the host DNA (maroon; flanking the other (IRL) end), which is bound non-specifically to the CC in a tract (yellow band) designated for target or host DNA. The reaction creates the branched figure-of-eight structure (precursor of the minicircle) with an interstitial sequence of host DNA (which will become the MCJ spacer between the abutted ends) equal in length to the distance between the two CCs. (B) In SC II, the two ends are complexed as in SC I with the MCJ spacer (black) spanning the distance between two active CCs. At each activated CC the 3' end of each IR is cleaved and the exposed 3'OH groups (broken strands with black half arrows) carry out concerted transesterification attacks (yellow dots) on target DNA (maroon) which is complexed through non-specific binding to the CCs (yellow tracts). This initiates the insertion event and the resulting direct repeats which are signatures of insertion will be equal in length to the MCJ spacer. BS: binding site; CC: catalytic center; CD: cleavage domain; IRR/IRL: right and left imperfect, inverted repeats; MCJ: minicircle junction; PBD: protein binding domain; SC: synaptic complex.