Direct Enzyme Engineering of B Family DNA Polymerases for Biotechnological Approaches.

Abstract

DNA-dependent DNA polymerases have been intensively studied for more than 60 years and underlie numerous biotechnological and diagnostic applications. In vitro, DNA polymerases are used for DNA manipulations, including cloning, PCR, site-directed mutagenesis, sequencing, and others. Understanding the mechanisms of action of DNA polymerases is important for the creation of new enzymes possessing improved or modified properties. This review is focused on archaeal family B DNA polymerases. These enzymes have high fidelity and thermal stability and are finding many applications in molecular biological methods. Nevertheless, the search for and construction of new DNA polymerases with altered properties is constantly underway, including enzymes for synthetic biology. This brief review describes advances in the development of family B DNA polymerases for PCR, synthesis of xeno-nucleic acids, and reverse transcription.