Direct expression of der f 2, a major house dust mite allergen, in escherichia i coli

Der f2 protein in a highly antigenic form was directly expressed in bacteria. Plasmid pFLU11 derived from pKK233-2 was designed to express methionyl-Der f 2 under the control of the trc promoter and it has the replication origin of pUC118 instead of its original to increase the copy number. This expression plasmid directed the synthesis of recombinant Der f 2 (rDer f 2) protein in an insoluble form of inclusion bodies in Escherichia coli cells. The high copy number plasmid pFLU11 conferred the efficient production of the Der f 2 protein in E. coli, when compared to a nonchanged origin material. rDer f 2 inclusion bodies were easily solubilized in urea and renatured by dialysis to assume the active form. The rDer f 2 protein was purified by means of anion exchange and gel filtration chromatography. This expression system yielded about 10 mg of purified rDer f 2 protein from the 1L culture. Purified rDer f 2 protein reacted with IgE from patient sera almost identically to the native Der f 2 in the RAST enzyme immunoassay and skin prick test.