Fig. 1
Crystal structures of RT-KTq/Mut_RT and Taq pol. (A) The overall crystal structure of RT-KTq and Mut_RT is shown as cartoon (adapted from PDB ID: 4BWM using PyMOL (Schrödinger, LLC, New York, NY)). Amino acid mutations specific for RT-KTq are coloured in blue and mutations specific for Mut_RT are coloured in green (side chains depicted as sticks). The KTq pol backbone is coloured in light grey. (B) The overall crystal structure of Taq pol is shown as cartoon (adapted from PDB ID: 1TAQ using PyMOL (Schrödinger, LLC, New York, NY)). The KTq pol backbone is coloured in light grey and the nuclease domain is depicted in dark grey. The amino acids that are changed to form the RT-Taq are coloured in blue. (C) Architecture of the KTq pol variants (top) and Taq pol (bottom). The positions of amino acid mutations are indicated in blue (RT-KTq, RT-Taq) and green (Mut_RT). (D) Combination strategy of the new Taq pol enzyme library. All eight amino acid mutations were combined to generate the wild-type Taq pol (without mutations), all single, double, triple and 4 − 8 fold mutants giving 256 Taq pol variants in total. The plasmid mix was transformed into E. coli cells, plated and 2660 single colonies were picked to form the library.
