Synthesis of preparative amounts of biologically active interleukin-6 using a continuous-flow cell-free translation system.
Cell-free synthesis of preparative amounts of interleukin-6 (IL-6) in a wheat germ extract is described. Following transcription of uncapped mRNA by SP6 polymerase, protein was synthesized in a continuous flow of translation components. The system remained active for at least 50 h, producing 1.25 x 10(6) units of IL-6. Recovered IL-6 was > 80% pure, reacted with anti-IL-6 polyclonal antiserum in Western blots, and had a specific activity comparable to recombinant IL-6 purified from Escherichia coli. This is the first time biologically active IL-6 has been prepared in preparative amounts using a continuous-flow cell-free translation system and confirms the feasibility of using enhanced in vitro translation for rapid synthesis of proteins from cloned templates.
Volyanik
E V
EV
Surgical Unit, London Hospital Medical College, University of London, Whitechapel, England.
Dalley
A
A
McKay
I A
IA
Leigh
I
I
Williams
N S
NS
Bustin
S A
SA
D000818:Animals; D001681:Biological Assay / Q000379:methods; D015153:Blotting, Western / Q000379:methods; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002474:Cell-Free System; D003001:Cloning, Molecular; D004926:Escherichia coli; D005813:Genes, Synthetic; D007106:Immune Sera; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000302:isolation & purification / Q000494:pharmacology; D007700:Kinetics; D051379:Mice; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000494:pharmacology; D012639:Seeds / Q000378:metabolism; D013698:Templates, Genetic; D014158:Transcription, Genetic; D014908:Triticum / Q000378:metabolism
D007106:Immune Sera; D015850:Interleukin-6; D012333:RNA, Messenger; D011994:Recombinant Proteins