Part:BBa_K4623003

TrwC, a protein linker to covalently link ssDNA 5' end

TrwC is a type of relaxase, which participates in bacterial DNA binding and horizontal transfer of genomic material between bacteria ^[1]. In vitro, it can specifically recognize the nic sequence (GGTGCGTATTGTCT^ATAGC) on ssDNA, causing it to break, and covalently bind to the ssDNA.

Therefore, TrwC relaxase possesses sequence specificity, orthogonality, and the ability to form a covalent bond with standard oligonucleotides. This provides a potential method in our project design that may be superior to existing protein-bound DNA nanostructures. The linkage of TrwC with ssDNA in vitro depends on Mg²⁺, so the reaction can be initiated by adding Mg²⁺ in the mixed system. Conversely, adding EDTA can terminate the reaction and yield DNA-protein complexes ^[2].

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal PstI site found at 286
Illegal PstI site found at 604
12
INCOMPATIBLE WITH RFC[12]
Illegal PstI site found at 286
Illegal PstI site found at 604
21
COMPATIBLE WITH RFC[21]
23
INCOMPATIBLE WITH RFC[23]
Illegal PstI site found at 286
Illegal PstI site found at 604
25
INCOMPATIBLE WITH RFC[25]
Illegal PstI site found at 286
Illegal PstI site found at 604
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 409
Illegal SapI.rc site found at 124

References:

[1]Lucas M, González-Pérez B, Cabezas M, Moncalian G, Rivas G, de la Cruz F. Relaxase DNA binding and cleavage are two distinguishable steps in conjugative DNA processing that involve different sequence elements of the nic site. J Biol Chem. 2010 Mar 19;285(12):8918-26. doi: 10.1074/jbc.M109.057539. Epub 2010 Jan 8. PMID: 20061574; PMCID: PMC2838313.

[2]Sagredo, Sandra et al. “Orthogonal Protein Assembly on DNA Nanostructures Using Relaxases.” *Angewandte Chemie (International ed. in English)* vol. 55,13 (2016): 4348-52. doi:10.1002/anie.201510313.