Immobilized proteins and use thereof

Abstract

The invention relates to an immobilized protein material comprising a protein that is immobilized on a glass material or organic polymer through affinity tag binding. The glass material may be a porous glass material such as (hybrid) controlled porosity glass. The invention also relates to the use of an immobilized enzyme material as a heterogeneous biocatalyst in chemical synthesis. The invention further relates to a method for the immobilization of affinity tagged proteins on a glass material or organic polymer, and to a method for the purification and isolation of affinity tagged proteins by the immobilization of such proteins on a glass material or organic polymer.

1 . A method for catalyzing an enzyme-catalyzed reaction, comprising
 (a) providing a carrier and at least one enzyme immobilized on the carrier, wherein the carrier comprises a porous organic polymer material to which an affinity matrix containing a chelated metal ion is attached,   
       wherein the at least one enzyme contains an affinity tag and is immobilized on the carrier through specific affinity binding to the affinity matrix; and
 (b) bringing the immobilized enzyme into contact with at least one substrate, thereby catalyzing the reaction. 
 
     
     
         2 . The method according to  claim 1 , wherein the affinity matrix is attached to the surface of the carrier through a linker. 
     
     
         3 . The method according to  claim 1 , wherein the porous organic polymer is chosen from the group consisting of polyethylene, ultra-high molecular weight polyethylene (UHMWPE), high-density polyethylene (HDPE), polypropylene (PP), polytetrafluoroethylene (PTFE), polyvinylidene fluoride (PVDF), polystyrene, polymethacrylate and poly(methyl methacrylate). 
     
     
         4 . The method according to  claim 1  wherein the porous organic polymer is a polystyrene. 
     
     
         5 . The method according to  claim 1 , wherein the porous organic polymer is a polymethacrylate. 
     
     
         6 . The method according to  claim 1 , wherein the porous organic polymer is present as porous particles with limited swelling. 
     
     
         7 . The method according to  claim 1 , wherein the at least one enzyme contains a polyhistidine tag and is immobilized on the carrier through specific affinity binding to the chelated metal ion. 
     
     
         8 . The method according to  claim 1 , wherein the chelated metal ion is Fe 2+ . 
     
     
         9 . The method according to  claim 1 , wherein the chelated metal ion is Fe 3+ . 
     
     
         10 . The method according to  claim 1 , wherein the chelated metal ion is Co 2+ . 
     
     
         11 . The method according to  claim 1 , wherein the chelated metal ion is Ni 2+ . 
     
     
         12 . The method according to  claim 1 , wherein the chelated metal ion is Cu 2+ . 
     
     
         13 . The method according to  claim 1 , wherein the chelated metal ion is Zn 2+ . 
     
     
         14 . The method according to  claim 1 , wherein two or more different enzymes are immobilized onto the carrier, wherein each of the different enzymes is able to catalyze a different reaction. 
     
     
         15 . The method according to  claim 1 , wherein the carrier additionally comprises metal nanoparticles. 
     
     
         16 . The method according to  claim 15 , wherein the said metal nanoparticles are transition metal nanoparticles. 
     
     
         17 . The method according to  claim 15 , wherein the metal nanoparticles are selected from the group consisting of cobalt, nickel and palladium nanoparticles. 
     
     
         18 . The method according to  claim 1 , wherein the enzyme-catalyzed reaction is performed under aqueous conditions. 
     
     
         19 . The method according to  claim 1 , wherein the enzyme-catalyzed reaction is performed in an organic solvent. 
     
     
         20 . The method according to  claim 1 , wherein the enzyme-catalyzed reaction is a continuous flow reaction. 
     
     
         21 . The method according to  claim 14 , wherein the enzyme-catalyzed reaction is a cascade reaction. 
     
     
         22 . The method according to  claim 21 , wherein a substrate for a first enzyme is transformed into a substrate for a second enzyme. 
     
     
         23 . The method according to  claim 21 , wherein a substrate is transformed by a first enzyme and wherein a co-factor for the first enzyme is regenerated by a second enzyme. 
     
     
         24 . The method according to  claim 16 , wherein a transition metal is immobilized on the carrier, and wherein the method is a multi-step reaction comprising an enzyme-catalyzed reaction and a transition metal-catalyzed reaction. 
     
     
         25 . The method according to  claim 1 , wherein the at least one substrate is converted to a product in the enzyme-catalyzed reaction.