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Characterisation of a DNA Polymerase Highly Mutated Along the Template Binding Interface

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Abstract

Phage display establishes a link between a polypeptide and its corresponding gene. It has been much used for the isolation of proteins binding to chosen molecular targets. A second link was designed more recently between a phage-displayed enzyme and its reaction product. Affinity chromatography for the product then allows the isolation of catalytically active enzymes and of their genes. Using this strategy, a polymerase with 15 mutations was selected by directed evolution of Thermus aquaticus DNA polymerase I. The kinetic characterisation reported here highlights the variant’s broad template specificity and classifies this enzyme as a thermostable DNA-dependent and RNA-dependent DNA-polymerase.

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Acknowledgements

This work was supported by the Ministère de la Recherche and by the Pasteur-Weizmann Council.

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Authors and Affiliations

  1. Département de Biologie Structurale et Chimie, Institut Pasteur, 28 rue du Dr. Roux, 75724, Paris 15, France

    Sophie Vichier-Guerre & Jean-Luc Jestin

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  1. Sophie Vichier-Guerre
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  2. Jean-Luc Jestin
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Correspondence to Jean-Luc Jestin.

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Vichier-Guerre, S., Jestin, JL. Characterisation of a DNA Polymerase Highly Mutated Along the Template Binding Interface. Mol Biotechnol 46, 58–62 (2010). https://doi.org/10.1007/s12033-010-9275-4

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