Disclosed herein are compositions for linking DNA-binding domains and cleavage domains to form nucleases, for example nucleases with altered target site separation (gap) preferences as compared to conventional linkers. Also described are fusion proteins comprising these linkers. The disclosure also provides methods of using these fusion proteins and compositions thereof for targeted cleavage of cellular chromatin in a region of interest and/or homologous recombination at a predetermined region of interest in cells. Thus, in one aspect, described herein are amino acid sequences linking a DNA-binding domain to a cleavage domain. The linker may replace one or more residues of the DNA-binding domain and/or cleavage domain.
