A method for the prevention of thrombin-induced degradation of recombinant proteins.
A new strategy to prevent degradation of recombinant proteins caused by non-specific cleavage by thrombin is described. We demonstrate that degradation due to non-specific cleavage of recombinant protein mediated by thrombin can be completely prevented by separation of thrombin from the recombinant protein on spin columns packed with heparin-sepharose. This method is generally applicable to all recombinant proteins that require the thrombin for the cleavage of affinity tags for purification. To our knowledge, this is the first report of an efficient and reliable method for the separation of residual thrombin from purified recombinant proteins.
Rajalingam
Dakshinamurthy
D
Department of Chemistry & Biochemistry, University of Arkansas, Fayetteville, AR 72701, USA.
Kathir
Karuppanan Muthusamy
KM
Ananthamurthy
Koteshwara
K
Adams
Paul D
PD
Kumar
Thallapuranam Krishnaswamy Suresh
TK
D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006493:Heparin / Q000737:chemistry; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012685:Sepharose / Q000737:chemistry; D013917:Thrombin / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*
D011994:Recombinant Proteins; D006493:Heparin; D012685:Sepharose; D013917:Thrombin