Expression and Characterization of Recombinant TGF-β2 Proteins Produced in Mammalian Cells

Recombinant DNA plasmids coding for transforming growth factor β2 (TGF-β2) precursor and a hybrid TGF-β1(NH2)/β2(COOH) molecule consisting of the amino-terminal precursor portion of transforming growth factor-β1 (TGF-β1) linked in phase to the carboxyl terminus of mature TGF-β2 were constructed and transfected into COS cells. Both plasmids directed the synthesis of active TGF-β2 which was secreted into the supernatants of transfected cells. The TGF-β2 was secreted in a latent form, as an acidification step was required to demonstrate optimal biological activity. Using site-specific anti-peptide antibodies, we show that precursor and mature forms of TGF-β2 are produced. A stable Chinese hamster ovary (CHO) cell line expressing the hybrid TGF-β1(NH2)/β2(COOH) protein was isolated. This cell line secreted both precursor and mature forms of TGF-β1(NH2)/β2(COOH); acidification was required to demonstrate biological activity. Protein sequence analysis of recombinant TGF-β2 produced by this CHO clone demonstrated that correct proteolytic cleavage had occurred, suggesting that the processing signals contained within the TGF-β1 amino portion can function in producing mature TGF-β2. Receptor binding studies showed that TGF-β2 specifically bound predominantly to type III receptors on the surface of human palatal mesenchymal cells. The availability of active TGF-β2 should aid in determining its potential therapeutic use as a growth modulator.