Microbial secretion of biologically active human transforming growth factor alpha fused to the Streptomyces protease inhibitor.
A secretory production system for the active form of transforming growth factor alpha (TGF alpha) was established in Streptomyces lividans using a gene encoding the secretory protease inhibitor, Streptomyces subtilisin inhibitor (SSI). It was demonstrated that deletion of one of the putative dual ssi terminators is effective to extracellularly produce a heterologous polypeptide in a fused form. The recombinant fusion protein, SSI::TGF alpha, was purified to homogeneity by a combination of hydrophobic chromatography and reverse-phase high-performance liquid chromatography (RP-HPLC). It was noteworthy that the SSI::TGF alpha hybrid protein exhibited bifunctional activity: the TGF alpha activity for cell growth promotion and the inhibitory activity of SSI. Taken together with the results of analytical gel filtration, these findings strongly indicate that each moiety in the fusion protein correctly folds and the whole hybrid molecule exists in a dimeric form, which results in its bifunctional activity.
Taguchi
S
S
Department of Biological Science and Technology, Science University of Tokyo, Chiba, Japan.
Misawa
S
S
Yoshida
Y
Y
Momose
H
H
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D018388:Codon, Terminator; D004305:Dose-Response Relationship, Drug; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D015842:Serine Proteinase Inhibitors / Q000235:genetics / Q000378:metabolism; D013302:Streptomyces / Q000235:genetics; D013381:Subtilisins / Q000037:antagonists & inhibitors; D016211:Transforming Growth Factor alpha / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology*
D001426:Bacterial Proteins; D018388:Codon, Terminator; D011993:Recombinant Fusion Proteins; D015842:Serine Proteinase Inhibitors; D016211:Transforming Growth Factor alpha; C018150:subtilisin inhibitor protein, Streptomyces; D013381:Subtilisins