Expression of human interleukin-11 cDNA in E.coli

【摘要】 <正> A 551-bp hIL-11 gene fragment that includes no nucleotide sequences encoding signalpolypeptide and the initial 8 amino acids of the mature protein was cloned into a high-level expression vectorpEx31B of E.coli.The authors identified the recombinant plasmid,designated pEx31-IL11,by restriction endonu-cleases digestion and DNA sequencing.The resulting recombinant plasmids were then used to transform E.colistrain HB101,and expression in the PL promoter system,which is temperature-regulated,was achieved.The ex-pressed fusion protein amounts to 50% of total bacterial proteins.The hIL-11 protein expressed in E.coliwas fused to the N-terminal 99 amino acids of the MS2 polymerase to form the inclusion body.Theserecombinant proteins can be purified to about 80% by extracting inclusion body with urea.OneIL-6-dependent cell line 7 TD1 was used for bioassay.The recombinant hIL-11 protein was preliminarily puri-fied and renatured to a specific activity of 10~5U/mg,even in the presence of an excess of a neutralizing an-ti-IL-6 antibody.更多还原

【Abstract】 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 endonu- cleases 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 ex- pressed 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 puri- fied and renatured to a specific activity of 10~5U/mg,even in the presence of an excess of a neutralizing an- ti-IL-6 antibody.更多还原