Expression of human interleukin-11 cDNA in E. coli.
A 551-bp hIL-11 gene fragment that includes no nucleotide sequences encoding signal polypeptide and the initial 8 amino acids of the mature protein was cloned into a high-level expression vector pEx31B of E. coli. The authors identified the recombinant plasmid, designated pEx31-IL11, by restriction endonucleases digestion and DNA sequencing. The resulting recombinant plasmids were then used to transform E. coli strain HB101, and expression in the PL promoter system, which is temperature-regulated, was achieved. The expressed fusion protein amounts to 50% of total bacterial proteins. The hIL-11 protein expressed in E. coli was fused to the N-terminal 99 amino acids of the MS2 polymerase to form the inclusion body. These recombinant proteins can be purified to about 80% by extracting inclusion body with urea. One IL-6-dependent cell line 7 TD1 was used for bioassay. The recombinant hIL-11 protein was preliminarily purified and renatured to a specific activity of 10(5)U/mg, even in the presence of an excess of a neutralizing anti-IL-6 antibody.
Miao
J
J
Institute of Basic Medical Sciences, Beijing, China.
Wang
J
J
Peng
S
S
Tang
P
P
Zou
M
M
Duan
J
J
Zhao
C
C
Ma
X
X
D018076:DNA, Complementary / Q000096:biosynthesis* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D017370:Interleukin-11 / Q000235:genetics*; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis*
D018076:DNA, Complementary; D017370:Interleukin-11; D011993:Recombinant Fusion Proteins