Reversible immobilization of arginine-tagged moieties on a silicate surface

Abstract

This invention provides materials and methods for the site specific attachment of virtually any moiety to a layered silicate surface. The methods involve covalently attaching the moiety to an arginine tag; and contacting the arginine tag with the layered silicate (e.g., mica).

1 - 33 . (canceled)  
     
     
         34 . A method of purifying a target molecule from a heterogeneous mixture of molecules, said method comprising the steps of: 
 a) providing a target molecule attached to an arginine tag; and    b) contacting said target molecule with a surface of a layered silicate; whereby said target molecule binds to said surface.    
     
     
         35 . The method of  claim 34 , wherein said arginine tag comprises at most two arginine residues.  
     
     
         36 . The method of  claim 34 , wherein said arginine tag comprises from about two to about one hundred residues.  
     
     
         37 . The method of  claim 34 , wherein said arginine tag consists of arginine residues.  
     
     
         38 . The method of  claim 34 , wherein said layered silicate is mica.  
     
     
         39 . The method of  claim 34 , further comprising contacting said mica surface with a sodium salt.  
     
     
         40 . The method of  claim 34 , wherein said contacting comprises flowing said heterogeneous mixture over one or more mica surfaces.  
     
     
         41 . The method of  claim 34 , wherein said contacting comprises combining said layered silicate with said heterogeneous mixture.  
     
     
         42 . The method of  claim 41 , wherein said method further comprises removing said layered silicate from said heterogeneous mixture.  
     
     
         43 . The method of  claim 42 , wherein said removing comprises centrifuging said heterogeneous mixture.  
     
     
         44 . The method of  claim 34 , further comprising the steps of contacting said layered silicate with a compound selected from the group consisting of a potassium salt, an arginine, and a poly-arginine, in a concentration sufficient to release said target molecule.  
     
     
         45 . The method of  claim 34 , wherein said target molecule is a fusion polypeptide comprising a polypeptide fused to said arginine tag at the carboxyl or the amino terminus of said arginine tag.  
     
     
         46 . The method of  claim 45 , wherein said fusion polypeptide further comprises a protease recognition site between said arginine tag and said target molecule.  
     
     
         47 . The method of  claim 45 , wherein said fusion polypeptide is recombinantly expressed.  
     
     
         48 . The method of  claim 34 , wherein said layered silicate is mica powder.  
     
     
         49 . The method of  claim 48 , wherein said mica powder is contained within a chromatography column.  
     
     
         50 . The method of  claim 34 , further comprising the steps of: 
 c) contacting said mica surface with a sodium salt; and    d) contacting said layered silicate surface with a compound selected from the group consisting of a potassium salt, an arginine, and a poly-arginine, in a concentration sufficient to release said target molecule.    
     
     
         51 - 55 . (canceled)  
     
     
         56 . The method of  claim 56 , wherein said protein is chosen from one or more of DNA binding protein, molecular motor, actin filament, microtubule, myosin filament, actin binding protein, and myosin filament binding protein.  
     
     
         57 . The method of  claim 56 , wherein said arginine tag comprises at least two arginine residues.  
     
     
         58 . The method of  claim 56 , wherein said arginine tag comprises from 2 to 100 arginine residues.  
     
     
         59 . The method of  claim 56 , wherein said layered silicate comprises mica.  
     
     
         60 . The method of  claim 56 , wherein said method further comprises contacting said surface of said layered silicate with a solution comprising a sodium salt in a concentration sufficient to remove molecules bound to said surface of said layered silicate by non-specific ion exchange.  
     
     
         61 . The method of  claim 60 , wherein said sodium salt is present in a concentration of at least 1 mM.  
     
     
         62 . The method of  claim 56 , wherein said layered silicate is chosen from one or more of vermiculite, montmorilonite, bentonite, hectorite, fluorohectorite, hydroxyl hectorite, muscovite boron fluorophlogopite, hydroxyl boron phlogopite, and mica.  
     
     
         63 . The method of  claim 56 , wherein said arginine tag comprises a homopolymer consisting of 6 contiguous arginine residues.