Activated formylglycine-generating enzymes and methods of producing and using the same
The present disclosure provides activated formylglycine-generating enzymes (FGE), methods of producing activated FGE, and their use in methods of producing a protein comprising a formylglycine (FGly) residue. The methods of producing activated FGE, as well as methods of use of activated FGE in producing FGly-containing proteins, include both cell-based and cell-free methods. Compositions and kits that find use, e.g., in practicing the methods of the present disclosure are also provided.
1. An activated formylglycine-generating enzyme (FGE) produced by a method comprising treating an FGE with Cu 2+ to produce an activated FGE.
2. A cell-free composition comprising:
an activated formylglycine-generating enzyme (FGE); and
a buffer.
3. The composition of claim 2 , further comprising a protein comprising an FGE recognition site.
4. The composition of claim 2 , wherein the FGE is an N-terminally truncated FGE.
5. The composition of claim 4 , wherein the FGE is a N-terminally truncated human FGE.
6. A kit comprising:
an activated formylglycine-generating enzyme (FGE); and
instructions for using the activated FGE to convert a cysteine residue or a serine residue present in an FGE recognition site of a protein to a formylglycine residue.
7. The kit of claim 6 , wherein the FGE is an N-terminally truncated FGE.
8. The kit of claim 7 , wherein the FGE is a N-terminally truncated human FGE.
9. A kit comprising:
a nucleic acid that encodes a formylglycine-generating enzyme (FGE); and
Cu 2+ or a source of Cu 2+ .
10. The kit of claim 9 , further comprising cells suitable for expressing the FGE encoded by the nucleic acid.