Method for purification of recombinant proteins
AssigneeClontech Laboratories, Inc.
InventorsPaul S. Nelson, Te-Tuan Yang, Steven R. Kain
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Abstract
Purification of poly-amino acid-tagged recombinant proteins has been improved by the use of a carboxymethylated aspartate ligand complexed with a third-block transition metal having an oxidation state of 2.sup.+ and a coordination number of 6. A method for synthesizing the metal ion-CM-Asp complex is also described. Further, the metal ion-CM-Asp complex can be used for screening protein function.
Claims6
An immobilized metal ion affinity chromatography purification method for purification of a recombinant protein, said method comprising:
- (a) providing carboxymethylated aspartate ligand complexed with a transition metal ion in a 2.sup.+ oxidation state, having a coordination number of 6;
- (b) loading a mixture of cell lysate comprising a recombinant protein having a polyhistidine tail to bind with said ligand:
- and
- (c) eluting said recombinant protein with a suitable elutant to obtain a purified recombinant protein, wherein said transition metal-complexed carboxymethylated aspartate ligand forms a carboxymethylated aspartate chelating matrix which comprises said transition metal and a polymer matrix, wherein said carboxymethylated aspartate chelating matrix has the structure ##STR2## wherein:
- M=transition metal ion in a 2.sup.+ oxidation state with a coordination number of 6;
- R.sub.1 =a linking arm connecting the nitrogen atom of CM-Asp with R.sub.2 ;
- R.sub.2 =a functional linking group through which CM-Asp linking arm R.sub.1 is connected to R.sub.3 ;
- and
- R.sub.3 =a polymer matrix.
An immobilized metal ion affinity chromatography complex, wherein said complex has the structure:
- ##STR3## wherein:
- M=transition metal ion in a 2.sup.+ oxidation state with a coordination number of 6;
- R.sub.1 =a linking arm connecting the nitrogen atom of CM-Asp with R.sub.2 ;
- R.sub.2 =a functional linking group through which CM-Asp linking arm R.sub.1 is connected to R.sub.3 ;
- and
- R.sub.3 =a polymer matrix.
Dependencies are inferred from the claim text.