PURIFICATION AND CHARACTERIZATION OF MICROBIALLY EXPRESSED NEOMYCIN PHOSPHOTRANSFERASE-II (NPTII) PROTEIN AND ITS EQUIVALENCE TO THE PLANT EXPRESSED PROTEIN

The gene encoding neomycin phosphotransferase II (NPTII) has been used routinely as a selectable marker in the production of genetically eng ineered crops. To facilitate the safety assessment of this protein, th e same coding sequence used for plant transformation was introduced in to Escherichia coli to produce gram quantities of this protein. A uniq ue, simple, rapid and efficient purification method was developed to p urify thirty grams of NPTII protein. The microbially produced NPTII wa s shown to be chemically and functionally equivalent to the NPTII prot ein expressed in and purified from genetically engineered cotton seed, potato tubers and tomato fruit. Microbially produced and plant produc ed NPTII proteins have comparable molecular weights, immuno-reactiviti es, epitope structures, amino terminal amino acid sequences, biologica l activities and both lack glycosylation. Demonstrating the equivalenc e of NPTII protein from these sources establishes the validity of usin g the microbially produced NPTII to assess the safety of the NPTII pro tein produced in genetically engineered crops.