Medical Research Council

United Kingdom Research and Innovation

Vitor B Pinheiro

Philipp Holliger

Christopher Cozens

Alexander I Taylor

Chris Cozens

The invention relates to a nucleic acid polymerase capable of producing a non-DNA nucleotide polymer from a DNA nucleotide template, said polymerase comprising amino acid sequence having at least 36% identity to the amino acid sequence of SEQ ID NO:1, wherein said amino acid sequence is mutated relative to the amino acid sequence of SEQ ID NO:1 at one or more residues of the thumb region, said residues selected from: amino acids 651 to 679 (patch 10A); wherein said amino acid sequence is mutated relative to the amino acid sequence of SEQ ID NO:1 at residue E664. In one embodiment said polymerase comprises the mutations Y409G and E664K. In one embodiment said polymerase comprises amino acid sequence corresponding to SEQ ID NO:12. The invention also relates to A nucleic acid polymerase capable of reverse transcribing a HNA nucleotide polymer into a DNA nucleotide polymer, said polymerase comprising amino acid sequence having at least 36% identity to the amino acid sequence of sEQ ID NO:1, wherein said amino acid sequence is mutated relative to the amino acid sequence of SEQ ID NO:1 at residue I521.

Enzymes

The invention relates to a nucleic acid polymerase capable of producing a non-DNA nucleotide polymer from a DNA nucleotide polymer template, the polymerase comprising amino acid sequence having at least 36% identity to the amino acid sequence of SEQ ID NO: 1, wherein the amino acid sequence is mutated relative to the amino acid sequence of SEQ ID NO: 1, wherein the amino acid sequence comprises the mutations P657T, E658Q, K659H, Y663H, D669A, K671N, and T676I; wherein the amino acid sequence is further mutated relative to the amino acid sequence of SEQ ID NO: 1 at residue: E664 and wherein the amino acid sequence comprises the mutation E664K. The invention also relates to methods of making nucleotide polymers comprising use of this polymerase. Suitably the nucleotides are arabino nucleotides such as ARA or FANA nucleotides.

Polymerase capable of producing non-DNA nucleotide polymers

The invention relates to a nucleic acid polymerase capable of producing a non-DNA nucleotide polymer from a DNA nucleotide polymer template, said polymerase comprising amino acid sequence having at least 36% identity to the amino acid sequence of SEQ ID NO:1, wherein said amino acid sequence is mutated relative to the amino acid sequence of SEQ ID NO:1 at one or more residues of the thumb region, said residues selected from: amino acids 651 to 679 (patch 10A); wherein said amino acid sequence is mutated relative to the amino acid sequence of SEQ ID NO:1 at residue E664. In one embodiment said polymerase comprises the mutations Y409G and E664K. In one embodiment said polymerase comprises amino acid sequence corresponding to SEQ ID NO:12. The invention also relates to A nucleic acid polymerase capable of reverse transcribing a HNA nucleotide polymer into a DNA nucleotide polymer, said polymerase comprising amino acid sequence having at least 36% identity to the amino acid sequence of SEQ ID NO:1, wherein said amino acid sequence is mutated relative to the amino acid sequence of SEQ ID NO:1 at residue I521.

The present invention relates to the development of a novel method for the selection of nucleic acid processing and other enzymes. In particular the invention relates to a method for the selection of nucleic acid polymerases and other enzymes with desired properties based on the method of compartmentalized self-tagging.

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Cambridge, United Kingdom

Vitor B Pinheiro