Expression of secreted insulin-like growth factor-1 in Escherichia coli.
The synthesis, processing and secretion of insulin-like growth factor-1 (IGF-1 or somatomedin-C) fused to LamB and OmpF secretion leader sequences in Escherichia coli have been investigated. Expression and secretion of IGF-1 was achieved. The major portion of this secreted IGF-1 accumulated in the periplasmic space as insoluble aggregates. A small amount of IGF-1 was found folded in its native conformation in the medium. The lamB and ompF signal sequences were fused to the 5' coding sequence of IGF-1. Fusion of the lamB signal sequence directly to IGF-1 (lamB-IGF-1) resulted in accumulation of 16-20 micrograms/A550/ml of correctly processed IGF-1 in the periplasmic space. The processing efficiency of LamB-IGF-1 and OmpF-IGF-1 was enhanced in an E. coli strain bearing a prlA4 mutation. Amino acid sequence analysis of IGF-1 secreted into the periplasm and exported into the medium confirmed the precise removal of the LamB or OmpF signal sequence. IGF-1 synthesized in E. coli was demonstrated to be active in a cell proliferation bioassay.
Wong
E Y
EY
Department of Biological Sciences, Monsanto Co., Chesterfield, MO 63198.
Seetharam
R
R
Kotts
C E
CE
Heeren
R A
RA
Klein
B K
BK
Braford
S R
SR
Mathis
K J
KJ
Bishop
B F
BF
Siegel
N R
NR
Smith
C E
CE
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development / Q000648:ultrastructure; D005796:Genes; D005813:Genes, Synthetic; D005822:Genetic Vectors; D007334:Insulin-Like Growth Factor I / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D015183:Restriction Mapping; D013002:Somatomedins / Q000235:genetics*
D011993:Recombinant Fusion Proteins; D013002:Somatomedins; D007334:Insulin-Like Growth Factor I