Recombinant human transforming growth factor-beta 1: expression by Chinese hamster ovary cells, isolation, and characterization.
A partial cDNA clone encoding the gene for human transforming growth factor-beta 1 (TGF-beta 1) was isolated from a human bladder carcinoma cell line (5637) cDNA library. Following restriction enzyme processing and ligation of synthetic oligonucleotide linkers, the gene was inserted into a plasmid and transfected into Chinese hamster ovary cells. Clonal selection and growth conditions resulted in a method for production of recombinant human TGF-beta 1 at 7 mg/liter in conditioned cell medium. Through a combination of low pH treatment, cation-exchange chromatography, and salt precipitation, the recombinant human TGF-beta 1 was purified in milligram amounts to > 95% purity in a yield of about 36%. Purification to homogeneity was accomplished by chromatography on C18 silica gel. Amino acid analysis, N-terminal sequencing, and growth inhibition assays indicate identity with the molecule from human platelets.
Bourdrel
L
L
Amgen, Inc., Amgen Center, Thousand Oaks, California 91320.
Lin
C H
CH
Lauren
S L
SL
Elmore
R H
RH
Sugarman
B J
BJ
Hu
S
S
Westcott
K R
KR
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001483:Base Sequence; D001681:Biological Assay / Q000379:methods; D016466:CHO Cells; D002455:Cell Division / Q000187:drug effects; D002461:Cell Line, Transformed; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D006224:Cricetinae; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D017421:Sequence Analysis; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification
D000596:Amino Acids; D003470:Culture Media; D016212:Transforming Growth Factor beta