Engineered gene-editing proteins

A method for gene-editing a cell, the method comprising:

• (a) providing a nucleic acid molecule encoding a gene-editing protein, the gene-editing protein comprising:
• (i) a DNA-binding domain comprising a plurality of repeat sequences and at least one of the repeat sequences comprising a repeat variable di-residue (RVD) at residue 12 or 13 which targets the DNA-binding domain to a target DNA molecule, wherein at least one of the repeat sequences comprises an amino acid sequence LTPvQVVAIAwxyzα (SEQ ID NO:
• 11) wherein v is E, w is S, x is I, H, N, or I, y is D, A, I, N, H, K, S, G, or null, z is SEQ ID NO:
• 8, and a is GTHG (SEQ ID NO:
• 62);
• and
• (ii) a nuclease domain comprising a catalytic domain of a nuclease which is capable of creating a single-strand break or a double-strand break in the target DNA molecule;
• wherein the nucleic acid molecule is a synthetic RNA molecule;
• and
• (b) contacting the cell with the synthetic RNA molecule to yield a single-strand break or a double-strand break in the target DNA molecule in the cell,
• wherein the contacting occurs at a temperature from about 30° C. to about 35° C.;
• and
• wherein the efficiency of the gene-editing at a temperature from about 30° C. to about 35° C. is greater than the efficiency of the gene-editing at a temperature of about 37° C.