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. 2013 Feb 1;288(11):7564–7571. doi: 10.1074/jbc.M112.434969

FIGURE 4.

FIGURE 4.

Acquirement of a higher RNA cleavage specificity in MazF-bs(can). A, analysis of cleavage sites in MS2 phage RNA by MazF-bs(arg) or MazF-bs(can). Lane C represents a control reaction in which no protein was added; MS2 RNA was incubated with MazF-bs(arg) or MazF-bs(can) for 1, 5, 10, and 30 min (lanes 1–4, MazF-bs(arg); lanes 5–8, MazF-bs(can)). A black arrow indicates a full-length (3.5 kb) of MS2 phage RNA. B–F, analysis of MazF-bs(can) cleavage sites in MS2 phage RNA by in vitro primer extension. Each panel represents different UACAU sites in MS2 RNA. Lane 1, MS2 RNA incubated with purified CspA. Lanes 2 and 3, MS2 RNA incubated with MazF-bs(can) and MazF-bs(arg) in the presence of CspA, an RNA chaperone, respectively. G, A, U, and C with an upper black bar indicate the sequence ladder for each reaction primer. Ribonucleotide sequences in each panel (B–F) indicate the cleavage sequences for MazF-bs(can) and MazF-bs(arg). The U↑ACAU sequences with an under bar indicate the cleavage sites by MazF-bs(arg); one extra ribonucleotide, A, at the 3′ end is colored in red, which is required for the cleavage by MazF-bs(can).