Expression and characterization of recombinant TGF-beta 2 proteins produced in mammalian cells.
Recombinant DNA plasmids coding for transforming growth factor beta 2 (TGF-beta 2) precursor and a hybrid TGF-beta 1(NH2)/beta 2(COOH) molecule consisting of the amino-terminal precursor portion of transforming growth factor-beta 1 (TGF-beta 1) linked in phase to the carboxyl terminus of mature TGF-beta 2 were constructed and transfected into COS cells. Both plasmids directed the synthesis of active TGF-beta 2 which was secreted into the supernatants of transfected cells. The TGF-beta 2 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-beta 2 are produced. A stable Chinese hamster ovary (CHO) cell line expressing the hybrid TGF-beta 1(NH2)/beta 2(COOH) protein was isolated. This cell line secreted both precursor and mature forms of TGF-beta 1(NH2)/beta 2(COOH); acidification was required to demonstrate biological activity. Protein sequence analysis of recombinant TGF-beta 2 produced by this CHO clone demonstrated that correct proteolytic cleavage had occurred, suggesting that the processing signals contained within the TGF-beta 1 amino portion can function in producing mature TGF-beta 2. Receptor binding studies showed that TGF-beta 2 specifically bound predominantly to type III receptors on the surface of human palatal mesenchymal cells. The availability of active TGF-beta 2 should aid in determining its potential therapeutic use as a growth modulator.
Madisen
L
L
Department of Entomology, Texas A&M University, College Station 77843.
Farrand
A L
AL
Lioubin
M N
MN
Marzowski
J
J
Knox
L B
LB
Webb
N R
NR
Lim
J
J
Purchio
A F
AF
D000818:Animals; D002460:Cell Line; D002999:Clone Cells; D006224:Cricetinae; D066246:ErbB Receptors / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D011506:Proteins / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics; D015291:Transforming Growth Factors / Q000032:analysis / Q000235:genetics*
D011506:Proteins; D011994:Recombinant Proteins; D015291:Transforming Growth Factors; D066246:ErbB Receptors