Figure 3: MR-B1H-produced zinc fingers provide high CCR5 activity with strong discrimination against CCR2.

(a) The corresponding CCR5 versus CCR2 sequences are shown with the 12-bp zinc-finger targets bold and underlined. Sequences are shown 5′ to 3′, but because ZFN monomers bind opposite strands of DNA, the left ZFN monomer targets the reverse complement of the sequence shown (compare with target in b). Mismatches with the CCR2 sequence are shown as red letters. (b) SELEX results for candidate zinc fingers. Candidates’ (Cand.) identification numbers and the sequences of their helices are listed above each SELEX plot. Helices that correspond to the discriminatory base are shown in red. The discriminatory base is boxed in green. The recovered percentage of sequences that correspond to the CCR5 or CCR2 base at this critical position are listed to the right of each plot. Those that tolerate binding to the selected-against base are shown as red numbers. The previously published helices for each left and right monomers are boxed at the top. (c) Candidate ZFN pairs were expressed in vivo and the percentage of indels at CCR5 and CCR2 measured (left and middle tables). The published candidates 8266–20505 were tested in parallel for reference. The ratio of CCR5 to CCR2 indel frequency is shown in the right table. The indel frequency recovered from a control that expressed GFP rather than a nuclease pair is shown below.