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Title: The Preprotein Binding Domain of SecA Displays Intrinsic Rotational Dynamics
Authors: Vandenberk, Niels
Karamanou, Spyridoula
Portaliou, Athina G.
Zorzini, Valentina
Hofkens, Johan
Hendrix, Jelle 
Economou, Anastassios
Issue Date: 2019
Publisher: CELL PRESS
Source: STRUCTURE, 27 (1), p. 90-101
Abstract: SecA converts ATP energy to protein translocation work. Together with the membrane-embedded SecY channel it forms the bacterial protein translocase. How secretory proteins bind to SecA and drive conformational cascades to promote their secretion remains unknown. To address this, we focus on the preprotein binding domain (PBD) of SecA. PBD crystalizes in three distinct states, swiveling around its narrow stem. Here, we examined whether PBD displays intrinsic dynamics in solution using single-molecule Forster resonance energy transfer (smFRET). Unique cysteinyl pairs on PBD and apposed domains were labeled with donor/acceptor dyes. Derivatives were analyzed using pulsed interleaved excitation and multi-parameter fluorescence detection. The PBD undergoes significant rotational motions, occupying at least three distinct states in dimeric and four in monomeric soluble SecA. Nucleotides do not affect smFRET-detectable PBD dynamics. These findings lay the foundations for single-molecule dissection of translocase mechanics and suggest models for possible PBD involvement during catalysis.
Notes: [Vandenberk, Niels; Hofkens, Johan; Hendrix, Jelle] Katholieke Univ Leuven, Dept Chem, Div Mol Imaging & Photon, Lab Photochem & Spect, Celestijnenlaan 200F, B-3001 Leuven, Belgium. [Karamanou, Spyridoula; Portaliou, Athina G.; Zorzini, Valentina; Economou, Anastassios] Katholieke Univ Leuven, Dept Microbiol & Immunol, Rega Inst Med Res, Lab Mol Bacteriol, Herestraat 49,Gasthuisberg Campus, B-3000 Leuven, Belgium. [Hendrix, Jelle] Hasselt Univ, Dynam Bioimaging Lab, Adv Opt Microscopy Ctr, Biomed Res Inst, Agoralaan C BIOMED, B-3590 Diepenbeek, Belgium.;
Keywords: single-molecule Förster resonance energy transfer; pulsed interleaved excitation; protein conformational dynamics; photon distribution analysis; bacterial secretion pathway; SecA motor ATPase; PBD domain motions
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ISSN: 0969-2126
e-ISSN: 1878-4186
DOI: 10.1016/j.str.2018.10.006
ISI #: 000454804900009
Category: A1
Type: Journal Contribution
Validations: ecoom 2020
Appears in Collections:Research publications

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