Covalent and selective immobilization of fusion proteins.
A general method for the covalent immobilization of fusion proteins is presented. The approach is based on the unusual mechanism of the human O6-alkylguanine-DNA alkyltransferase, which irreversibly transfers the alkyl group from its substrate, alkylated or benzylated guanine, to a reactive cysteine residue. By attaching the benzyl group to a surface, hAGT fusion proteins immobilize themselves in a specific and covalent manner. The specificity of the reaction of hAGT with its substrate even allows the specific immobilization of hAGT fusion proteins directly out of cell extracts, making the approach an attractive alternative to currently used immobilization procedures.
Kindermann
Maik
M
Institute of Molecular and Biological Chemistry, Swiss Federal Institute of Technology, CH-1015 Lausanne, Switzerland.
George
Nathalie
N
Johnsson
Nils
N
Johnsson
Kai
K
D015374:Biosensing Techniques / Q000379:methods; D005982:Glutathione Transferase / Q000032:analysis / Q000737:chemistry; D006147:Guanine / Q000031:analogs & derivatives* / Q000737:chemistry; D007700:Kinetics; D009154:Mutation; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000737:chemistry* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry*; D020349:Surface Plasmon Resonance
D011993:Recombinant Fusion Proteins; C064976:O(6)-benzylguanine; D006147:Guanine; D019853:O(6)-Methylguanine-DNA Methyltransferase; D005982:Glutathione Transferase