Methods for removing suspended particles from soluble protein solutions

Abstract

The present invention provides soluble protein solutions, free of suspended particles in high yield. More particularly, the current invention provides a method for removing suspended particles from soluble protein solutions by filtering the soluble protein solution through highly purified diatomaceous earth.

1 - 23 . (canceled)  
     
     
         24 . A method for removing suspended particles from a soluble protein solution comprising the step of filtering the soluble protein solution through highly purified diatomaceous earth, thereby providing a clarified soluble protein solution.  
     
     
         25 . The method of  claim 24 , wherein the soluble protein solution is a secreted protein solution.  
     
     
         26 . The method of  claim 25 , wherein the soluble protein solution is a lysate.  
     
     
         27 . The method of  claim 26 , wherein the lysate is a bacterial lysate.  
     
     
         28 . The method of  claim 26 , wherein the lysate is a bacterial lysate containing a heterologous protein that was obtained by expression in bacteria.  
     
     
         29 . The method of  claim 24 , further comprising the step of stirring the soluble protein solution with a highly purified diatomaceous earth before filtering through a filter press.  
     
     
         30 . The method of  claim 24 , wherein the yield of the soluble protein solution is between about 95% and about 100%.  
     
     
         31 . The method of  claim 24 , wherein the highly purified diatomaceous earth is CELPURE.  
     
     
         32 . The method of  claim 27 , wherein the bacteria is  E. coli.    
     
     
         33 . The method of  claim 28 , further comprising the step of blocking cysteine residues of the heterologous protein.  
     
     
         34 . The method of  claim 33 , wherein the cysteine residues are blocked with an oxidizing agent.  
     
     
         35 . The method of  claim 34 , wherein the oxidizing agent comprises sodium sulfite.  
     
     
         36 . The method of  claim 34 , wherein the oxidizing agent comprises sodium tetrathionate.  
     
     
         37 . The method of  claim 34 , wherein the oxidizing agent is a 2:1 ratio mixture of sodium sulfite and sodium tetrathionate.  
     
     
         38 . The method of  claim 34 , wherein the oxidizing agent is added to the protein solution at a pH of between about 7.8 and about 8.2.  
     
     
         39 . The method of  claim 33 , further comprising the step of deblocking the blocked cysteine residues.  
     
     
         40 . The method of  claim 39 , wherein the blocked cysteine residues are deblocked with a reducing agent.  
     
     
         41 . The method of  claim 39 , wherein the blocked cysteine residues are deblocked with dithiothreitol.  
     
     
         42 . The method of  claim 28 , further comprising resolubilizing refractile bodies in the lysate.  
     
     
         43 . The method of  claim 28 , in which the heterologous protein is SY161, wherein SY161 has an amino acid sequence as shown in SEQ ID NO 1.