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. 2015 Jun 22;43(13):6631–6648. doi: 10.1093/nar/gkv612

Figure 6.

Figure 6.

Extein–intein interactions affect temperature dependence of Pho RadA intein splicing. (A) Kinetics of splicing of extein mutants M1, M6 and M8. Mutants are compared with the wild-type FL-RadAi precursor (WT) at 55°C. Proteins were overexpressed in BL21 Star (DE3) cells at 46°C and Ni-NTA purified. The experiment was performed in triplicate and error bars represent standard deviation. All splicing assays have very small deviations, obscuring error bars in the plot. Initial rates of splicing are shown on the right. * indicates that the initial rate for M1 was calculated for the sample that had 50% of the precursor converted in vivo and therefore is likely to be an underestimate. (B) Splicing of extein mutant M1. WT and M1 mutant proteins were overexpressed in ArcticExpress (DE3) cells at 12°C. Mutant M1 and the wild-type FL-RadAi precursor (WT) were compared at 55, 45 and 35°C. All splicing assays have very small deviations, obscuring error bars in the plot. The experiment was performed in triplicate and error bars represent standard deviation. Initial rates of splicing at different temperatures are shown on the right.