Development of a Recombinant Bacterial Expression System for the Active Form of a Human Transforming Growth Factor β Type II Receptor Ligand Binding Domain

Assembly of TβRI:TβRII:TGFβ Ternary Complex in vitro with Receptor Extracellular Domains is Cooperative and Isoform-dependent

“…TbRII-ED has been expressed both fused to thioredoxin 25 and alone. 30,31 The fusion protein remained soluble and yielded active protein, 25 while TbRII-ED expressed alone was insoluble but could be refolded to yield active protein. 30,31 To prepare TbRI-ED, we tested the two approaches described above.…”

Mitochondrial Fatty Acid Synthesis in Trypanosoma brucei

“…Expression was induced with 0.1 mM isopropyl 1-thio-␤-D-galactopyranoside for 4 -5 h at 37°C. Virtually all of the recombinant ACP was sequestered in inclusion bodies, 3 which were purified essentially as described (20). Briefly, they were suspended in a buffer containing 2 M urea and 0.1% Triton X-100, washed several times, and then stored in 4 mM dithiothreitol at Ϫ80°C.…”

An efficient method for expression in Escherichia coli and purification of the extracellular ligand binding domain of the human TGFβ type II receptor

“…Moreover Asp118 and Glu119 were characterized as interfacial amino acid residues in the T␤RII-ED-TGF␤3 complex and probably were critical for binding (Hart et al, 2002). N-Terminal truncated T␤RII-ED (residues 22-136 or 27-136) without any affinity tags have been expressed in E. coli DH5a cells and refolded from inclusion bodies to an active form with yield of 5 mg protein from 1 L cell culture (Boesen et al, 2000). The fusion glutathione S-transferase-T␤RII-ED was expressed in soluble form, cleaved by thrombin and T␤RII-ED was purified at a yield of 10 g of T␤RII-ED/L of bacterial growth in the same work.…”

“…Unglycosylated T␤RII-ED produced by E. coli demonstrates a higher anti-TGF␤1 activity than T␤RII-ED produced by P. pastoris (Glansbeek et al, 1998). Expression and purification of soluble T␤RII-ED in E. coli by refolding of insoluble protein from inclusion bodies have been described earlier (Boesen et al, 2000). In this study T␤RII-ED expression and purification yielded 1-2 mg product from 1 L cell culture.…”