Method of labeling sulfenic acid-containing proteins and peptides

Abstract

A method of labeling a sulfenic acid (—SOH) group of a cysteine residue in a protein or peptide, comprises contacting said protein or peptide with a beta-ketoester to covalently couple said beta-ketoester to said cysteine residue and form a beta-ketoester-labeled cysteine residue in said protein or peptide.

That which is claimed is: 
     
         1 . A method of labeling a sulfenic acid (—SOH) group of a cysteine residue in a protein or peptide, comprising:
 contacting said protein or peptide with a beta-ketoester to covalently couple said beta-ketoester to said cysteine residue and form a beta-ketoester-labeled cysteine residue in said protein or peptide. 
 
     
     
         2 . The method of  claim 1 , further comprising the step of:
 detecting said beta-ketoester-labeled cysteine residue in said protein or peptide.   
     
     
         3 . The method of  claim 1 , further comprising the step of coupling a detectable group, or a protein or peptide binding ligand, to said beta-ketoester. 
     
     
         4 . The method of  claim 3 , further comprising the step of detecting said detectable group. 
     
     
         5 . The method of  claim 1 , further comprising the step of cleaving said beta-ketoester-labeled cysteine residue to produce a 5-isoxazolone-labeled cysteine residue in said protein or peptide. 
     
     
         6 . The method of  claim 5 , further comprising the step of detecting said 5-isoxazolone-labeled cysteine residue in said protein or peptide. 
     
     
         7 . The method of  claim 1 , wherein said contacting step is carried by contacting said beta-ketoester to a live cell containing said protein or peptide. 
     
     
         8 . The method of  claim 1 , wherein said protein or peptide is selected from the group consisting of enzymes, receptors, ion channels, transcription factors, hormones, receptor ligands, and enzyme substrates. 
     
     
         9 . The method of  claim 1 , wherein said beta-ketoester is a compound of Formula I: 
       
         
           
           
               
               
           
         
       
       wherein:
 X is H, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, cycloalkylalkenyl, cycloalkylalkynyl, heterocyclo, heterocycloalkyl, heterocycloalkenyl, heterocycloalkynyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, heteroaryl, heteroarylalkyl, heteroarylalkenyl, heteroarylalkynyl, alkoxy, halo, mercapto, azido, cyano, formyl, carboxylic acid, hydroxyl, nitro, acyl, aryloxy, alkylthio, amino, alkylamino, arylalkylamino, disubstituted amino, acylamino, acyloxy, ester, amide, sulfoxyl, sulfonyl, sulfonate, sulfonic acid, sulfonamide, urea, alkoxylacylamino, aminoacyloxy, or a detectable group; 
 Y is a covalent bond or an alkylene group or a linker; and 
 Z is H, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, cycloalkylalkenyl, cycloalkylalkynyl, heterocyclo, heterocycloalkyl, heterocycloalkenyl, heterocycloalkynyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, heteroaryl, heteroarylalkyl, heteroarylalkenyl, heteroarylalkynyl, alkoxy, halo, mercapto, azido, cyano, formyl, carboxylic acid, hydroxyl, nitro, acyl, aryloxy, alkylthio, amino, alkylamino, arylalkylamino, disubstituted amino, acylamino, acyloxy, ester, amide, sulfoxyl, sulfonyl, sulfonate, sulfonic acid, sulfonamide, urea, alkoxylacylamino, and aminoacyloxy, a detectable group, or a reactive group. 
 
     
     
         10 . A beta-ketoester for use in covalently labeling an oxidized sulfenic acid cysteine residue of a protein or peptide. 
     
     
         11 . The beta-ketoester of  claim 10 , wherein said protein or peptide is selected from the group consisting of enzymes, receptors, ion channels, transcription factors, hormones, receptor ligands, and enzyme substrates. 
     
     
         12 . The beta-ketoester of  claim 10 , wherein said beta-ketoester is a compound of Formula I: 
       
         
           
           
               
               
           
         
       
       wherein:
 X is H, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, cycloalkylalkenyl, cycloalkylalkynyl, heterocyclo, heterocycloalkyl, heterocycloalkenyl, heterocycloalkynyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, heteroaryl, heteroarylalkyl, heteroarylalkenyl, heteroarylalkynyl, alkoxy, halo, mercapto, azido, cyano, formyl, carboxylic acid, hydroxyl, nitro, acyl, aryloxy, alkylthio, amino, alkylamino, arylalkylamino, disubstituted amino, acylamino, acyloxy, ester, amide, sulfoxyl, sulfonyl, sulfonate, sulfonic acid, sulfonamide, urea, alkoxylacylamino, aminoacyloxy, or a detectable group; 
 Y is a covalent bond or an alkylene group or a linker; and 
 Z is H, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, cycloalkylalkenyl, cycloalkylalkynyl, heterocyclo, heterocycloalkyl, heterocycloalkenyl, heterocycloalkynyl, aryl, arylalkyl, arylalkenyl, arylalkynyl, heteroaryl, heteroarylalkyl, heteroarylalkenyl, heteroarylalkynyl, alkoxy, halo, mercapto, azido, cyano, formyl, carboxylic acid, hydroxyl, nitro, acyl, aryloxy, alkylthio, amino, alkylamino, arylalkylamino, disubstituted amino, acylamino, acyloxy, ester, amide, sulfoxyl, sulfonyl, sulfonate, sulfonic acid, sulfonamide, urea, alkoxylacylamino, and aminoacyloxy, a detectable group, or a reactive group.