Cyclopropene amino acids and methods

1. A polypeptide comprising an amino acid having a cyclopropene group wherein said cyclopropene group is linked to the amino acid via a carbamate group and the carbamate group does not form a part of the polypeptide backbone, wherein said cyclopropene group is a 1,3-disubstituted cyclopropene.

2. A polypeptide according to claim 1 wherein said cyclopropene is a 1,3-dimethylcyclopropene.

3. A polypeptide according to claim 1 wherein said cyclopropene group is present as a residue of a lysine amino acid.

4. A polypeptide according to claim 1 further comprising a tetrazine compound linked to said cyclopropene group.

5. An amino acid comprising cyclopropene wherein said cyclopropene group is linked to the amino acid moiety via a carbamate group and the carbamate group does not form a part of the peptide backbone, wherein said cyclopropene group is a 1,3-disubstituted cyclopropene.

6. An amino acid according to claim 5 wherein said cyclopropene is a 1,3-dimethylcyclopropene.

7. An amino acid according to claim 5 wherein said amino acid is a lysine amino acid.

8. An amino acid according to claim 7 which comprises N ε -[((2-methylcycloprop-2-en-1-yl)methoxy)carbonyl]-1-lysine.

9. An amino acid according to claim 7 which consists of

10. A method of producing a polypeptide comprising a cyclopropene group wherein said cyclopropene group is joined to an amino acid moiety of the polypeptide via a carbamate group, said method comprising genetically incorporating said amino acid comprising said cyclopropene group joined to said amino acid moiety via said carbamate group, into said polypeptide.

11. A method according to claim 10 wherein producing the polypeptide comprises

(i) providing a nucleic acid encoding the polypeptide which nucleic acid comprises an orthogonal codon encoding the amino acid having a cyclopropene group;

(ii) translating said nucleic acid in the presence of an orthogonal tRNA synthetase/tRNA pair capable of recognising said orthogonal codon and incorporating said amino acid having a cyclopropene group into the polypeptide chain.

12. A method according to claim 10 wherein said orthogonal codon comprises an amber codon (TAG), said tRNA comprises MbtRNA CUA and said tRNA synthetase comprises MbPylRS; or wherein said orthogonal codon comprises an amber codon (TAG), said tRNA comprises MmtRNA CUA and said tRNA synthetase comprises MmPylRS.

13. A method according to claim 10 wherein said carbamate group does not form a part of the peptide backbone, and wherein said cyclopropene group is a 1,3-disubstituted cyclopropene.

14. A method of producing a polypeptide comprising a tetrazine group, said method comprising providing a polypeptide according to claim 1 , contacting said polypeptide with a tetrazine compound, and incubating to allow joining of the tetrazine to a cyclopropene group of the polypeptide by an inverse electron demand Diels-Alder cycloaddition reaction.

15. A method according to claim 14 wherein said reaction is allowed to proceed for 10 minutes or less, preferably for 1 minute or less, preferably for 30 seconds or less.

16. A polypeptide according to claim 1 wherein said polypeptide comprises two or more amino acids each having a cyclopropene group, wherein each said cyclopropene group is linked to each said amino acid via a carbamate group and wherein each carbamate group does not form a part of the polypeptide backbone.

17. A polypeptide according to claim 16 wherein said polypeptide comprises four amino acids each having a cyclopropene group.

18. An antibody drug conjugate (ADC) comprising a polypeptide according to claim 1 .