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Sci Rep. 2015 Jul 27;5:12470. doi: 10.1038/srep12470.

Directionality of substrate translocation of the hemolysin A Type I secretion system.

Author information

1
Institute of Biochemistry, Heinrich-Heine-Universitaet, D-40225 Duesseldorf, Germany.
2
Center for Advanced Imaging (CAi), Heinrich-Heine-Universitaet, D-40225 Duesseldorf, Germany.
3
Protein Production Facility, Heinrich-Heine-Universitaet, 40225 Duesseldorf, Germany.
4
1] Institute of Molecular Enzyme Technology, Heinrich-Heine-Universitaet and Institute of Bio- and Geosciences IBG-1: Biotechnology, Forschungszentrum Juelich GmbH, D-52426 Juelich, Germany [2] Center of Excellence on Plant Sciences (CEPLAS), Heinrich-Heine-Universitaet, D-40225 Duesseldorf, Germany.
5
1] Institute of Biochemistry, Heinrich-Heine-Universitaet, D-40225 Duesseldorf, Germany [2] Center of Excellence on Plant Sciences (CEPLAS), Heinrich-Heine-Universitaet, D-40225 Duesseldorf, Germany.

Abstract

Type 1 secretion systems (T1SS) of Gram-negative bacteria are responsible for the secretion of various proteases, lipases, S-layer proteins or toxins into the extracellular space. The paradigm of these systems is the hemolysin A (HlyA) T1SS of Escherichia coli. This multiple membrane protein complex is able to secrete the toxin HlyA in one step across both E. coli membranes. Common to all secreted T1SS substrates is a C-terminal secretion sequence being necessary as well as sufficient for secretion. However, it is not known whether transport occurs directionally, i.e. the N- or the C-terminus of T1SS substrates is secreted first. We have addressed this question by constructing HlyA fusions with the rapidly folding eGFP resulting in a stalled T1SS. Differential labeling and subsequent fluorescence microscopic detection of C- and N-terminal parts of the fusions allowed us to demonstrate vectorial transport of HlyA through the T1SS with the C-terminus appearing first outside the bacterial cells.

PMID:
26212107
PMCID:
PMC4648471
DOI:
10.1038/srep12470
[Indexed for MEDLINE]
Free PMC Article

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