Compounds and methods for selective c-terminal labeling

Patent metadata

Field	Value
Publication number	US-11358981-B2
Application number	US-202117153490-A
Country	US
Kind code	B2
Filing date	Jan 20, 2021
Priority date	Jan 21, 2020
Publication date	Jun 14, 2022
Grant date	Jun 14, 2022

Abstract

The present disclosure relates to compounds and methods for selective C-terminal functionalization of peptides. In certain embodiments, the compounds have improved water-solubility, and are suitable for use in connection with peptide sequencing methodologies.

First claim

The invention claimed is:

1. A method of selective N-functionalization of a peptide, comprising reacting a plurality of peptides of Formula (XI):

or a salt thereof, wherein each P independently is a peptide having an N-terminal amine, with a compound of Formula (XII):

under conditions, comprising Cu 2± , or a precursor thereof, and a buffer having a pH of 10-11; to obtain a plurality of ε-azido compounds of the Formula (XIIIb):

or a salt thereof.

2. The method of claim 1 , wherein the plurality of peptides of Formula (XI), or a salt thereof, is obtained by subjecting a protein to enzymatic digestion to obtain a digestive mixture comprising the plurality of peptides of Formula (XI), or a salt thereof.

3. The method of claim 1 , further comprising reacting the plurality of compounds of Formula (XIIIb), or a salt thereof, with a compound of Formula (XIV):
R 6 -L 5 -Z 2    (XIV)

wherein R 6 is a moiety comprising an alkyne or a strained alkene;
L 5 is a linker or is absent; and
Z 2 comprises one or more of polyethylene glycol, single-stranded DNA, double-stranded DNA, biotin, and streptavidin;
to obtain a plurality of compounds of Formula (XV):

or a salt thereof, wherein Y 2 is a moiety resulting from a click reaction with the azide moiety of Formula (XIIIb), or a salt thereof, and R 6 .

4. The method of claim 3 , wherein Z 2 comprises single-stranded DNA.

5. The method of claim 2 , wherein the enzymatic digestion comprises cleaving the C-terminal bonds of lysine and/or arginine residues of the protein.

6. The method of claim 2 , wherein the protein comprises sulfide moieties, and wherein the sulfide moieties of the protein are protected prior to the enzymatic digestion.

7. The method of claim 3 , wherein Z 2 comprises double-stranded DNA.

8. The method of claim 3 , wherein Z 2 comprises biotin.

9. The method of claim 8 , wherein the biotin is bis-biotin (BisBt).

10. The method of claim 3 , wherein Z 2 comprises streptavidin.

11. The method of claim 3 , wherein R 6 comprises an alkyne.

12. The method of claim 11 , wherein the alkyne is bicycle[6.1.0]nonyne (BCN) or dibenzocyclooctyne (DBCO).

13. The method of claim 3 , wherein R 6 comprises an alkene.

14. The method of claim 13 , wherein the akene is trans-cyclooctene (TCO).

15. The method of claim 3 , wherein L 5 is absent.

16. The method of claim 1 , wherein N-terminal:εselectivity is at least about 90%.

Assignees

• Quantum Si Inc

Inventors

• Huang Haidong
• Nani Roger
• Ad Omer
• Boer Robert E

Classifications

• C07K1/34

  by filtration, ultrafiltration or reverse osmosis · CPC title

• C07K1/12

  by hydrolysis {, i.e. solvolysis in general} · CPC title

• C07K1/1077 Primary

  by covalent attachment of residues other than amino acids or peptide residues, e.g. sugars, polyols, fatty acids · CPC title

• C07K1/128 Primary

  sequencing · CPC title

• C07K14/195

  from bacteria · CPC title