Synthesis of Site Specifically-Linked Ubiquitin

摘要

The invention relates to a method of modifying a specific lysine residue in a polypeptide comprising at least two lysine residues, said method comprising
(a) providing a polypeptide comprising a target lysine residue protected by a first protecting group, and at least one further lysine residue;(b) treating the polypeptide to protect said further lysine residue(s), wherein the protecting group for said further lysine residues is different to the protecting group for the target lysine residue;(c) selectively deprotecting the target lysine residue; and(d) modifying the deprotected lysine residue of (c).

主权项

1. A method of modifying a specific lysine residue in a ubiquitin polypeptide comprising at least two lysine residues, said method comprising:
(a) providing a ubiquitin polypeptide comprising a target lysine residue protected by a first protecting group, and at least one further lysine residue, wherein the ubiquitin polypeptide comprising the target lysine group is provided by genetically incorporating the target lysine residue protected by the first protecting group into the ubiquitin polypeptide chain during its translation by
(i) providing a nucleic acid encoding the ubiquitin polypeptide which nucleic acid comprises an orthogonal codon encoding the target lysine; and(ii) translating said nucleic acid in the presence of an orthogonal tRNA synthetase/tRNA pair capable of recognising said orthogonal codon and incorporating said target lysine residue protected by a first protecting group into the ubiquitin polypeptide chain; (b) treating the ubiquitin polypeptide to protect said further lysine residue(s), wherein the protecting group for said further lysine residues is different to the protecting group for the target lysine residue; (c) selectively deprotecting the target lysine residue; and (d) modifying the deprotected lysine residue of (c).