2005
DOI: 10.1007/s00018-005-5165-5
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Directed evolution as a tool for understanding and optimizing nucleic acid polymerase function

Abstract: Polynucleotide polymerases play a crucial role in transmitting genetic information from generation to generation, and they are the most important reagents in biotechnology. Although classical crystal structure analyses as well as biochemical studies have significantly contributed to our understanding of how DNA polymerases function, surprising new insights regarding the importance of certain residues and protein motifs, or of their mutability have been achieved in recent years by evolutionary approaches. Direc… Show more

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Cited by 18 publications
(4 citation statements)
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References 86 publications
(104 reference statements)
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“…Yet for several applications the characteristics of natural evolved DNA polymerases are not satisfying. Thus, several modified DNA polymerases with new characteristics have been developed by directed evolution, a promising method to create nucleic acid polymerases with modified properties. Nevertheless, as noncanonical amino acids in general could be also very promising tools for DNA polymerase engineering, our studies may open up new opportunities for investigations like the directed evolution of DNA polymerases with new characteristics due to the expanded amino acid repertoire. …”
Section: Discussionmentioning
confidence: 99%
“…Yet for several applications the characteristics of natural evolved DNA polymerases are not satisfying. Thus, several modified DNA polymerases with new characteristics have been developed by directed evolution, a promising method to create nucleic acid polymerases with modified properties. Nevertheless, as noncanonical amino acids in general could be also very promising tools for DNA polymerase engineering, our studies may open up new opportunities for investigations like the directed evolution of DNA polymerases with new characteristics due to the expanded amino acid repertoire. …”
Section: Discussionmentioning
confidence: 99%
“…Mutations of the DNA polymerase itself, by site-specific or random mutagenesis, are effective ways to create modified enzymes with improved PCR performance or specific properties for in vitro DNA manipulations. An artificial evolution procedure also has attracted a great deal of attention, for the creation of DNA polymerases with novel activities (Brakmann, 2005; Henry and Romesberg, 2005; Holmberg et al, 2005; Ong et al, 2006). Our strategy of using environmental DNA as a genetic resource also works well to investigate the structure–function relationships of DNA polymerases.…”
Section: Future Perspectivesmentioning
confidence: 99%
“…First, mutations are introduced throughout the whole gene or in selected regions resulting in a library of enzyme variants. [27,28] After creation of an enzyme library, an expression and separation process is required. That step has to involve a connection between the specific enzyme genotype and its respective phenotype.…”
Section: Directed Dna Polymerase Evolutionmentioning
confidence: 99%