Directed evolution of mesophilic DNA polymerases with altered substrate specificity

Many parts of biology are universal: present in every organism, made by the same processes, based on the same chemistry and following the same rules. Rules that emerged very early in evolution such as the genetic code and the storage of genetic information in DNA and RNA. Nevertheless, that universality does not imply that biology has explored every possible chemical
reaction or every possible permutation. Thus, if a viable alternative exists, it should be possible to implement it through biology – with the engineering challenge depending on how big a change is
being implemented. Here we set out to change the chemistry of the enzymes responsible for propagating DNA, called polymerases. Polymerases make use of reactive building blocks for making DNA in a cell or in a test tube. Those building blocks are universal but other chemical alternatives exist – building blocks that can be used for making DNA but that cannot be used by natural enzymes. Using molecular evolution, our goal is to engineer a natural polymerase to synthesize DNA with the modified building blocks and become unable to use the natural ones. We will tackle the engineering problem by molecular evolution as well as the knowledge we will acquire by testing many enzyme variants. This is part of a wider program focusing on developing improved containment of genetically modified organisms.