Chimeric polypeptides having targeted binding specificity
Disclosed herein are chimeric polypeptides, including compositions thereof, expression vectors, and methods of use thereof, for the generation of transgenic cells, tissues, plants, and animals. The compositions, vectors, and methods of the present invention are also useful in gene therapy techniques.
1 . A polypeptide comprising a Ralstonia derived transcription activator-like effector (TALE) protein, the TALE protein having an N-terminal domain (NTD) comprising an amino acid sequence as set forth in SEQ ID NO: 8 (IVDIAR1QR2SGDLA) having one or more mutations or deletions selected from: R1 is K, Q is Y, Q is S, Q is R, R2 is W, R2 is G, R2 is deleted, S is R, Sis H, Sis A, S is N, and Sis T.
2 . The polypeptide of claim 1 , wherein the NTD comprises an amino acid sequence selected from: IVDIARQWSGDLA (SEQ ID NO: 9), IVDIARYRGDLA (SEQ ID NO: 10), IVDIARSRSGDLA (SEQ ID NO: 11), IVDIARYHGDLA (SEQ ID NO: 12), and IVDIARRGAGDLA (SEQ ID NO: 13).
3 . The polypeptide of claim 1 , wherein the TALE protein further comprises a synthetic repeat variable di-residues (RVD) domain.
4 . The polypeptide of claim 1 , wherein the TALE protein comprises a C-terminal or N-terminal truncation relative to a wildtype Ralstonia TALE protein from which the TALE protein was derived.
5 . The polypeptide of claim 1 , further comprising a recombinase domain or a nuclease domain.
6 . The polypeptide of claim 5 , wherein the recombinase domain is selected from the group consisting of Gin, Hin, Tn3, Sin, Beta, Pin, Min, Din, Cin, muteins of Gin, muteins of Hin, muteins of Tn3, muteins of Sin, muteins of Beta, muteins of Pin, muteins of Min, muteins of Din, and muteins of Cin.
7 . A nucleic acid molecule encoding the polypeptide of claim 1 .
8 . An expression cassette comprising the nucleic acid molecule of claim 7 .
9 . A vector comprising the expression cassette of claim 8 .
10 . A host cell transformed or transfected with the vector of claim 9 .