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. 2010 Mar 17;132(14):5096–5104. doi: 10.1021/ja909180c

Figure 10.

Figure 10

To determine the relative phenotypic contributions of the selected mutations of Pfu E10 (V337I, E399D, N400G, R407I, Y546H), we determined activities of either wild-type Pfuexo- polymerase or Pfu variants comprising only A-motif mutations (A: Pfuexo- E399D, N400G, R407I)) or C-motif mutations (C: Pfuexo- Y546H) in (a) polymerase ELISA with Cy3- or Cy5-dCTP and (b) PCR with unmodified dNTPs. Both (a) Cy3- and Cy5-dCTP polymerase-ELISA and (b) PCR reveal that there is a strong synergistic epistasis between A- and C-motif mutations for the incorporation of Cy3- or Cy5-dCTP, which also results in an unexpected shortening of extension times in standard PCR compared to wild-type Pfuexo- polymerase.