Orientation and density control of proteins on solid matters by outer membrane coating: Analytical and diagnostic applications


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Abstract

HighlightsA lack of orientation control exists when solid matters are coated with biomolecules.This leads to a severe reduction in sensitivity and the limit of detection.Autodisplay leads to an oriented expression of proteins on the OM.OM liposomes can be prepared and used for solid matter coating.Orientation and density control obtained thereby leads to an increase in sensitivity and LOD.Autodisplay is an expression system for the display of recombinant proteins on the outer membrane (OM) of gram negative bacteria and has been developed for translocation studies, whole cell biocatalysis, bioremediation, inhibitor screening, and enzyme refolding. Recently, affinity proteins such as IgG-binding Z-domains and biotin-binding streptavidin have been autodisplayed on the OM of Escherichia coli for analytical and biomedical applications. The secretion mechanism of the autodisplay system was used and orientation and density control of these affinity proteins were determined. Affinity protein-autodisplaying E. coli cells have been used to coat solid supports in immunoassays. For this purpose, the OM of autodisplayed E. coli cells was separated and isolated by the aid of detergents. The structure of the resulting OM liposomes as well as their physico-chemical parameters, were analyzed. OM liposomes were used subsequently for coating various solid matters including microplates and biosensor transducer surfaces and the formation of OM layers were monitored. OM layer formation on solid matters was shown to increase the sensitivity of immunoassays and biosensors. In this review, analytical and diagnostic applications are described in particular concerning orientation and density control of autodisplayed affinity proteins.

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