Membrane translocation assay based on proteolytic cleavage: application to diphtheria toxin T domain.

ABSTRACT: The function of diphtheria toxin translocation (T) domain is to transfer the catalytic domain across the endosomal membrane upon acidification. The goal of this study was to develop and apply an in vitro functional assay for T domain activity, suitable for investigation of structure-function relationships of translocation across lipid bilayers of various compositions. Traditionally, T domain activity in vitro is estimated by measuring either conductance in planar lipid bilayers or the release of fluorescent markers from lipid vesicles. While an in vivo cell death assay is the most relevant to physiological function, it cannot be applied to studying the effects of pH or membrane lipid composition on translocation. Here we suggest an assay based on cleavage of the N-terminal part of T domain

SUBMITTER: Rodnin MV 

PROVIDER: S-EPMC4259893 | biostudies-literature | 2015 Jan

REPOSITORIES: biostudies-literature