Secretion of Human Angiogenin into Periplasm and Culture Medium with Its Eukaryotic Signal Sequence by Escherichia coli

  • Jung, Woo-Jung (Department of Chemistry, Kangnung National University) ;
  • Choi, Suk-Jung (Department of Chemistry, Kangnung National University)
  • Received : 1996.10.18
  • Published : 1997.01.31

Abstract

The synthesis and secretion of human angiogenin in E. coli by the natural leader sequence has been studied. We constructed a recombinant plasmid containing human angiogenin cDNA which encompassed all the coding region including leader sequence required for secretion. The recombinant plasmid was introduced into a suitable E. coli host. The angiogenin was detected in the culture medium and periplasm upon the induction of gene expression. The molecular weight of the secreted angiogenin was identical to that of authentic angiogenin purfied from human plasma when estimated by SDS-PAGE and immunoblotting. showing that the natural leader sequence was recognized and processed by the secretion machinery of E. coli. The angiogenin concentration in the culture medium reached a maximum within 2 h when expressed at $37^{\circ}C$ with 0.02~2 mM IPTG. In contrast, the expression level increased gradually over time up to 11 h at $23^{\circ}C$ with 0.002~2 mM IPTG and at $37^{\circ}C$ with 0.002 mM IPTG.

Keywords

eukaryotic leader sequence;Escherichia coli;human angiogenin;secretory expression

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