Detecting protein analytes that modulate transmembrane movement of a polymer chain within a single protein pore

Liviu Movileanu, Stefan Howorka, Orit Braha, Hagan Bay-ley

Research output: Contribution to journalArticle

256 Scopus citations

Abstract

Here we describe a new type of biosensor element for detecting proteins in solution at nanomolar concentrations. We tethered a 3.4 kDa polyethylene glycol chain at a defined site within the lumen of the transmembrane protein pore formed by staphylococcal α-hemolysin. The free end of the polymer was covalently attached to a biotin molecule. On incorporation of the modified pore into a lipid bilayer, the biotinyl group moves from one side of the membrane to the other, and is detected by reversible capture with a mutant streptavidin. The capture events are observed as changes in ionic current passing through single pores in planar bilayers. Accordingly, the modified pore allows detection of a protein analyte at the single-molecule level, facilitating both quantification and identification through a distinctive current signature. The approach has higher time resolution compared with other kinetic measurements, such as those obtained by surface plasmon resonance.

Original languageEnglish (US)
Pages (from-to)1091-1095
Number of pages5
JournalNature Biotechnology
Volume18
Issue number10 SUPPL.
DOIs
StatePublished - Jan 1 2000
Externally publishedYes

Keywords

  • Biosensor
  • Nanostructure
  • Polymer
  • Pore
  • Protein engineering

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Molecular Medicine
  • Biomedical Engineering

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