SPLIT INTEINS, CONJUGATES AND USES THEREOF
Disclosed herein are split inteins, fused proteins of split inteins, and methods of using split inteins to efficiently purify and modify proteins of interest. Thus, provided herein are fusion proteins of a polypeptide and a split intein N-fragment, or variant thereof, as described below in greater detail. Also provided are complexes of the fusion protein and a split intein C-fragment or variant thereof as described in detail below. The complex of the fusion protein and C-fragment or variant thereof can be via a covalent interaction between the fusion protein and C-fragment or variant or via a noncovalent interaction (e.g., ionic, H-bonding, and/or van der Waals interaction). Further provided herein are split intein C-fragments or variants thereof.
1 . A fusion protein comprising a split intein N-fragment and a polypeptide, wherein the split intein N-fragment comprises a sequence selected from the group consisting of SEQ ID NOs: 1-19, or a variant thereof.
2 . The fusion protein of claim 1 , wherein the split intein N-fragment comprises a sequence of SEQ ID NO: 19, or a variant thereof.
3 . The fusion protein of claim 1 , wherein the split intein N-fragment comprises a sequence selected from the group consisting of SEQ ID NO: 20-38 and 761, or a variant thereof.
4 . The fusion protein of claim 1 , wherein the polypeptide has a molecular weight of 40 kDa or greater.
5 . The fusion protein of claim 4 , wherein the polypeptide is an antibody or fragment thereof.
6 . The fusion protein of claim 5 wherein the antibody is an IgG antibody or fragment thereof.
7 . The fusion protein of claim 6 , wherein a heavy chain of the antibody is fused to the split intein N-fragment.
8 . The fusion protein of claim 7 , wherein each heavy chain of the antibody is fused to the split intein N-fragment.
9 . The fusion protein of claim 6 , wherein a light chain of the antibody is fused to the split intein N-fragment.
10 . The fusion protein of claim 1 , wherein the polypeptide is secreted from a cell.
11 . A complex comprising the fusion protein of claim 1 and the split intein C-fragment of claim 12 .
12 . A split intein C-fragment comprising a sequence selected from the group consisting of SEQ ID NOs: 57-74, or a variant thereof.
13 . The split intein C-fragment of claim 12 comprising a sequence selected from the group consisting of SEQ ID NOs: 75-128, or a variant thereof.
14 . The split intein C-fragment of claim 12 comprising a sequence of SEQ ID NO: 707, or a variant thereof.
15 . The split intein C-fragment of claim 14 comprising a sequence selected from the group consisting of SEQ ID NOs: 708-711, or a variant thereof.
16 . The split intein C-fragment of claim 12 attached to a support.
17 . The split intein C-fragment of claim 16 , wherein the support comprises a particle, a bead, a resin, or a slide.
18 . The split intein C-fragment of claim 17 , wherein the split intein C-fragment comprises a sequence selected from the group consisting of SEQ ID NOs: 417-704 and 745-759, or a variant thereof.
19 . A method comprising
(a) contacting (1) a fusion protein comprising a polypeptide and a split intein N-fragment comprising a sequence selected from the group consisting of SEQ ID NO: 1-19, or a variant thereof, and (2) a split intein C-fragment comprising a sequence selected from the group consisting of SEQ ID NO: 57-74, or a variant thereof, wherein contacting is performed under conditions that permit binding of the split intein N-fragment to the split intein C-fragment to form an intein intermediate; and
(b) contacting the intein intermediate with a nucleophile to form a conjugate of the protein and the nucleophile.
20 - 34 . (canceled)