Assessing the efficacy of vesicle fusion with planar membrane arrays using a mitochondrial porin as reporter.

Autor: Pszon-Bartosz K; Aquaporin A/S, Ole Maaløes Vej 3, DK-2200 Copenhagen N, Denmark., Hansen JS, Stibius KB, Groth JS, Emnéus J, Geschke O, Hélix-Nielsen C
Jazyk: angličtina
Zdroj: Biochemical and biophysical research communications [Biochem Biophys Res Commun] 2011 Mar 04; Vol. 406 (1), pp. 96-100. Date of Electronic Publication: 2011 Feb 03.
DOI: 10.1016/j.bbrc.2011.02.001
Abstrakt: Reconstitution of functionally active membrane protein into artificially made lipid bilayers is a challenge that must be overcome to create a membrane-based biomimetic sensor and separation device. In this study we address the efficacy of proteoliposome fusion with planar membrane arrays. We establish a protein incorporation efficacy assay using the major non-specific porin of Fusobacterium nucleatum (FomA) as reporter. We use electrical conductance measurements and fluorescence microscopy to characterize proteoliposome fusion with an array of planar membranes. We show that protein reconstitution in biomimetic membrane arrays may be quantified using the developed FomA assay. Specifically, we show that FomA vesicles are inherently fusigenic. Optimal FomA incorporation is obtained with a proteoliposome lipid-to-protein molar ratio (LPR)=50 more than 10(5) FomA proteins could be incorporated in a bilayer array with a total membrane area of 2mm(2) within 20 min. This novel assay for quantifying protein delivery into lipid bilayers may be a useful tool in developing biomimetic membrane applications.
(Copyright © 2011 Elsevier Inc. All rights reserved.)
Databáze: MEDLINE
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