One-step reduction and alkylation of proteins

Abstract

The present invention relates generally to the field of analyzing proteins and more specifically to the reduction and alkylation of protein samples during protein analysis. A novel composition and method for reducing and alkylating proteins is disclosed. A novel reagent including a combination of a volatile reducing agent, a volatile alkylating agent, and a volatile solvent is used for a one-step reduction and alkylation of proteins allowing the protein sample to remain in the same container during the reduction and alkylation processes.

1 . A composition for one-step reduction and alkylation of proteins, comprising: 
 a substantially volatile reducing agent;    a substantially volatile solvent; and    a substantially volatile and water soluble alkylating agent.    
   
   
       2 . The composition of  claim 1 , wherein the composition substantially dissolves the protein at solvent compositions having greater than about 30% water.  
   
   
       3 . The composition of  claim 1 , wherein the composition substantially dissolves the protein at solvent compositions having greater than about 50% water.  
   
   
       4 . The composition of  claim 1 , wherein the alkylating agent comprises a halogenated alcohol.  
   
   
       5 . The composition of  claim 1 , wherein the volatile solvent comprises acetonitrile.  
   
   
       6 . The composition of  claim 1 , wherein the alkylating agent is selected from the group consisting of iodoethanol, bromoethanol, iodopropanol, bromopropanol, iodomethanol, bromomethanol and mixtures thereof.  
   
   
       7 . The composition of  claim 1 , wherein the concentration of the alkylating agent is between about 0.1% to about 1.0% by volume of the composition.  
   
   
       8 . The composition of  claim 1 , wherein the reducing agent is selected from the group consisting of triethylphosphine, trimethylphosphine, tripropylphosphine, tributylphosphine and mixtures thereof.  
   
   
       9 . The composition of  claim 8 , wherein the concentration of the reducing agent is between about 0.025% to about 0.25% by volume of the composition.  
   
   
       10 . The composition of  claim 1 , wherein the solvent includes water and acetonitrile.  
   
   
       11 . The composition of  claim 1 , wherein the volatile solvent is up to 50% water by volume, based on the total volume of the composition.  
   
   
       12 . The composition of  claim 1 , wherein the volatile reagent includes a buffer configured to buffer the pH of the composition to about pH 10.  
   
   
       13 . The composition of  claim 12 , wherein the buffer is 0.1M ammonium carbonate.  
   
   
       14 . A composition for one-step reduction and alkylation of a protein sample, comprising: 
 a reducing agent to reduce the protein sample;    an alkylating agent to alkylate the protein sample, the alkylating agent being water soluble;    a solvent to substantially dissolve the protein sample; and    wherein the composition is substantially volatile, and further wherein the composition dissolves the protein at solvent compositions of greater than about 30% water.    
   
   
       15 . The composition of  claim 14 , wherein the composition dissolves the protein at solvent compositions of greater than about 50% water.  
   
   
       16 . The composition of  claim 14 , wherein the alkylating agent comprises a halogenated alcohol.  
   
   
       17 . The composition of  claim 14 , wherein the alkylating agent is selected from the group consisting of iodoethanol, bromoethanol, iodopropanol, bromopropanol, iodomethanol, bromomethanol and mixtures thereof.  
   
   
       18 . The composition of  claim 14 , wherein the concentration of the alkylating agent is between about 0.1% to about 1.0% of the composition by volume.  
   
   
       19 . The composition of  claim 14 , wherein the reducing agent is selected from the group consisting of triethylphosphine, trimethylphosphine, tripropylphosphine, tributylphosphine and mixtures thereof.  
   
   
       20 . The composition of  claim 19 , wherein the concentration of the reducing agent is between about 0.025% to about 0.25% by volume of the composition.