Methods and compositions for genome engineering

1. A zinc finger protein that binds to an endogenous human albumin gene, the zinc finger protein comprising five or six zinc finger domains designated and ordered F1 to F5 or F1 to F6, the zinc finger domains comprising the recognition helix regions ordered and shown as follows:

(i) F1: QSGNLSR (SEQ ID NO:23);

F2: LKQNLCM (SEQ ID NO:18);

F3: WADNLQN (SEQ ID NO:24);

F4: TSGNLTR (SEQ ID NO:20); and

F5: RQSHLCL (SEQ ID NO:21), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:17; or

(ii) F1: TPQLLDR (SEQ ID NO:14);

F2: LKWNLRT (SEQ ID NO:9);

F3: DQSNLNA (SEQ ID NO:25);

F4: RNFSLTM (SEQ ID NO:15);

F5: LRHDLDR (SEQ ID NO:16); and

F6: HRSNLNK (SEQ ID NO:12), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:7; or

(iii) F1: QSGNLAR (SEQ ID NO:2);

F2: LIQYLQS (SEQ ID NO:26);

F3: WADNLQN (SEQ ID NO:24);

F4: TSGNLTR (SEQ ID NO:20); and

F5: RQSHLSL (SEQ ID NO:27), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:17; or

(iv) F1: TPQLLDR (SEQ ID NO:14);

F2: LKWNLRT (SEQ ID NO:9);

F3: DQSNLRA (SEQ ID NO:10);

F4: RNFSLTM (SEQ ID NO:15);

F5: LRHDLDR (SEQ ID NO:16); and

F6: HRSNLNK (SEQ ID NO:12), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:7; or

(v) F1: QSGNLAR (SEQ ID NO:2);

F2: LIQYLQS (SEQ ID NO:26);

F3: WADNLQN (SEQ ID NO:24);

F4: TSGNLTR (SEQ ID NO:20); and

F5: RQSHLCL (SEQ ID NO:21), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:17; or

(vi) F1: TPQLLDR (SEQ ID NO:14);

F2: LKHNLLT (SEQ ID NO:28);

F3: DQSNLNA (SEQ ID NO:25);

F4: RNFSLTM (SEQ ID NO:15);

F5: LRHDLDR (SEQ ID NO:16); and

F6: HRSNLNK (SEQ ID NO:12), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:7; or

(vii) F1: TPQLLDR (SEQ ID NO:14);

F2: LKWNLRT (SEQ ID NO:9);

F3: DQSNLRA (SEQ ID NO:10);

F4: RNFSLTM (SEQ ID NO:15);

F5: LRHDLDR (SEQ ID NO:16); and

F6: HRSNLNK (SEQ ID NO:12), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:7; or

(viii) F1: QSGNLAR (SEQ ID NO:2);

F2: LIQYLQS (SEQ ID NO:26);

F3: WQSNLQN (SEQ ID NO:19);

F4: TSGNLTR (SEQ ID NO:20); and

F5: RQSHLCL (SEQ ID NO:21), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:17; or

(ix) F1: QSSDLSR (SEQ ID NO:8);

F2: LKHNLLT (SEQ ID NO:28);

F3: LKHNLLT (SEQ ID NO:28);

F4: RPYTLRL (SEQ ID NO:11);

F5: LRPDLER (SEQ ID NO:41); or

F6: HRSNLNK (SEQ ID NO:12), wherein the zinc finger protein binds to a target site as shown in SEQ ID NO:7.

2. A fusion protein comprising a zinc finger protein of claim 1 and a wild-type or engineered cleavage domain or cleavage half-domain.

3. A polynucleotide encoding one or more proteins of claim 1 .

4. An expression vector comprising a polynucleotide according to claim 3 .

5. The expression vector of claim 4 , wherein the vector is an AAV vector.

6. The expression vector of claim 5 , wherein the AAV vector is an AAV2/8 vector.

7. A pharmaceutical composition comprising an expression vector according to claim 4 .

8. A method of cleaving an albumin gene in a cell, the method comprising: introducing, into the cell, one or more expression vectors comprising a polynucleotide which encodes a fusion protein of claim 2 , under conditions such that the one or more proteins are expressed and the albumin gene is cleaved.

9. The method of claim 8 , further comprising introducing a donor sequence into the cell such that the donor sequence is integrated into the cleaved albumin gene.

10. The method of claim 9 , wherein the donor sequence is introduced using an AAV vector.

11. The method of claim 10 , wherein the AAV vector is an AAV2/8 vector.

12. The method of claim 9 , wherein the donor sequence encodes a protein lacking or deficient in a lysosomal storage disease (LSD).

13. The method of claim 8 , wherein the cell is a liver cell.

14. A kit comprising an expression vector according to claim 4 .