title: Directionality of substrate translocation of the hemolysin A Type I secretion system.
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.
authors.affiliation: Institute of Biochemistry, Heinrich-Heine-Universitaet, D-40225 Duesseldorf, Germany.
authors.forename: Michael H H
authors.initials: MH
authors.lastname: Lenders
authors.affiliation: Center for Advanced Imaging (CAi), Heinrich-Heine-Universitaet, D-40225 Duesseldorf, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Weidtkamp-Peters
authors.affiliation: Protein Production Facility, Heinrich-Heine-Universitaet, 40225 Duesseldorf, Germany.
authors.forename: Diana
authors.initials: D
authors.lastname: Kleinschrodt
authors.affiliation: 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.
authors.forename: Karl-Erich
authors.initials: KE
authors.lastname: Jaeger
authors.affiliation: Institute of Biochemistry, Heinrich-Heine-Universitaet, D-40225 Duesseldorf, Germany.
authors.forename: Sander H J
authors.initials: SH
authors.lastname: Smits
authors.affiliation: 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.
authors.forename: Lutz
authors.initials: L
authors.lastname: Schmitt
mesh_terms: D004926:Escherichia coli / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000378:metabolism*; D000069284:Protein Translocation Systems / Q000502:physiology*; D021381:Protein Transport / Q000502:physiology*
chemical_list: D029968:Escherichia coli Proteins; D006460:Hemolysin Proteins; C036507:Hlya protein, E coli; D000069284:Protein Translocation Systems