Fig 2. Triphosphorylation kinetics of central ribozyme variants in this study.
The starting point of the doped selection was the ribozyme TPR1 (empty circles). It has a kobs of 0.013 min-1 under selection conditions (100 mM MgCl2, 50 mM trimetaphosphate, 50 mM Tris/HCl pH 8.3). The most efficient isolate from the doped selection was a 16-mutation variant called clone 11 (filled triangles, kobs of 0.21 min-1). After removal of unnecessary mutations a 5-mutation variant called TPR1-II resulted (open squares, kobs of 0.25 min-1). Two mutations that arose independently were introduced to yield TPR1e (filled squares), a 7-mutation variant with a kobs of 0.31 min-1. Lines are single-exponential curve fits to the data. Error bars denote the standard deviations from triplicate experiments.