Efficient cleavage by alpha-thrombin of a recombinant fused protein which contains insulin-like growth factor I.
The gene for insulin-like growth factor I (IGF-I) was constructed from chemically synthesized deoxyoligonucleotides and expressed in Escherichia coli, under the control of a trp promoter, as a set of fusion proteins which were connected with a portion of human growth hormone through the recognition sequence for a sequence-specific protease, either blood coagulation factor Xa or alpha-thrombin. Upon induction with 3-indoleacrylic acid, fusion proteins accumulated with a yield of 10-30% of the total protein. A fusion protein connected through a tetradecapeptide (Asp-Asp-Pro-Pro-Thr-Val-Glu-Leu-Gln-Gly-Leu-Val-Pro-Arg) was efficiently and correctly cleaved by alpha-thrombin, and the purified IGF-I possessed somatomedin-like activity, as determined by the enhancement of sulfation of glycosaminoglycans in cultured costal chondrocytes from rabbits.
Nishikawa
S
S
Faculty of Pharmaceutical Sciences, Osaka University, Japan.
Yanase
K
K
Tokunaga-Doi
T
T
Kodama
K
K
Gomi
H
H
Uesugi
S
S
Ohtsuka
E
E
Kato
Y
Y
Suzuki
F
F
Ikehara
M
M
D001681:Biological Assay; D011088:DNA Ligases / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D015951:Factor Xa; D005786:Gene Expression Regulation; D005813:Genes, Synthetic; D007334:Insulin-Like Growth Factor I / Q000235:genetics / Q000378:metabolism*; D010447:Peptide Hydrolases / Q000378:metabolism; D010957:Plasmids; D011863:Radioimmunoassay; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D012697:Serine Endopeptidases / Q000378:metabolism; D013002:Somatomedins / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D013002:Somatomedins; D007334:Insulin-Like Growth Factor I; D010447:Peptide Hydrolases; D012697:Serine Endopeptidases; D013917:Thrombin; D015951:Factor Xa; D011088:DNA Ligases