Isolation and characterization of the outer membrane of Escherichia coli with autodisplayed Z-domains

Min Park, Gu Yoo, Ji Hong Bong, Joachim Jose, Min Jung Kang, Jae Chul Pyun

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

"Autodisplay technology" is an expression technique used to display the various recombinant proteins on the outer membrane (OM) of Escherichia coli. The resulting autodisplayed Z-domain has been used to improve the sensitivity of immunoassays. In this work, a facile isolation method of the OM fraction of E. Coli with autodisplayed Z-domains was presented using (1) an enzyme reaction for the hydrolysis of the peptidoglycan layer and (2) short centrifugation steps. The purity of the isolated OM fraction was analyzed. For the estimation of contamination with bacterial proteins from other parts of E. Coli, Western blots of marker proteins for the OM (OmpA), periplasm (β-lactamase), inner membrane (SecA), and cytoplasm (β-galactosidase) were performed. Additionally, assays of marker components or enzymes from each part of E. Coli were carried out including the OM (KDO), inner membrane (NADH oxidase), periplasm (β-lactamase), and cytoplasm (β-galactosidase). The yield of OM isolation using this new method was determined to be 80% of the total OM amount, with less than 1% being contaminants from other parts of E. Coli.

Original languageEnglish
Pages (from-to)842-847
Number of pages6
JournalBiochimica et Biophysica Acta - Biomembranes
Volume1848
Issue number3
DOIs
Publication statusPublished - 2015 Jan 15

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Escherichia coli
Membranes
Galactosidases
Periplasm
Cytoplasm
Bacterial Proteins
Centrifugation
Peptidoglycan
Enzymes
Immunoassay
Recombinant Proteins
Hydrolysis
Assays
Contamination
Western Blotting
Impurities
Technology

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Cell Biology

Cite this

Park, Min ; Yoo, Gu ; Bong, Ji Hong ; Jose, Joachim ; Kang, Min Jung ; Pyun, Jae Chul. / Isolation and characterization of the outer membrane of Escherichia coli with autodisplayed Z-domains. In: Biochimica et Biophysica Acta - Biomembranes. 2015 ; Vol. 1848, No. 3. pp. 842-847.
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Isolation and characterization of the outer membrane of Escherichia coli with autodisplayed Z-domains. / Park, Min; Yoo, Gu; Bong, Ji Hong; Jose, Joachim; Kang, Min Jung; Pyun, Jae Chul.

In: Biochimica et Biophysica Acta - Biomembranes, Vol. 1848, No. 3, 15.01.2015, p. 842-847.

Research output: Contribution to journalArticle

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