Structure and membrane-targeting of a Bordetella pertussis effector N-terminal domain

Adi Yahalom; Geula Davidov; Sofiya Kolusheva; Hadassa Shaked; Shiran Barber-Zucker; Raz Zarivach; Jordan H. Chill

doi:10.1016/j.bbamem.2019.183054

Structure and membrane-targeting of a Bordetella pertussis effector N-terminal domain

Adi Yahalom, Geula Davidov, Sofiya Kolusheva, Hadassa Shaked, Shiran Barber-Zucker, Raz Zarivach, Jordan H. Chill

Ben-Gurion University of the Negev

Research output: Contribution to journal › Article › peer-review

9 Scopus citations

Abstract

BteA, a 69-kDa cytotoxic protein, is a type III secretion system (T3SS) effector in the classical Bordetella, the etiological agents of pertussis and related mammalian respiratory diseases. Like other cytotoxicity-mediating effectors, BteA uses its multifunctional N-terminal domain to target phosphatidylinositol (PI)-rich microdomains in the host membrane. Despite their structural similarity, T3SS effectors exhibit a variable range of membrane interaction modes, and currently only limited structural information is available for the BteA membrane-targeting domain and the molecular mechanisms underlying its function. Employing a synergistic combination of structural methods, here we determine the structure of this functional domain and uncover key molecular determinants mediating its interaction with membranes. Residues 29–121 of BteA form an elongated four-helix bundle packed against two shorter perpendicular helices, the second of which caps the domain in a critical ‘tip motif’. A flexible region preceding the BteA helical bundle contains the characteristic β-motif required for binding its cognate chaperone BtcA. We show that BteA targets PI(4,5)P₂-containing lipoprotein nanodiscs and binds a soluble PI(4,5)P₂ analog via an extensive positively charged surface spanning its first two helices, and that this interaction is weaker for PI(3,5)P₂ and abolished for PI(4)P. We confirmed this model of membrane-targeting by observation of BteA-induced changes in the structure of PI(4,5)P₂-containing phospholipid bilayers using small-angle X-ray scattering (SAXS). We also extended these results to a larger BteA domain (residues 1–287), confirming its interaction with bilayers using calorimetry, fluorescence and SAXS methods. This novel view of the structural underpinnings of membrane targeting by BteA is an important step towards a comprehensive understanding of cytotoxicity in Bordetella, as well as interactions of a broad range of pathogens with their respective hosts.

Original language	English
Article number	183054
Journal	Biochimica et Biophysica Acta - Biomembranes
Volume	1861
Issue number	12
DOIs	https://doi.org/10.1016/j.bbamem.2019.183054
State	Published - 1 Dec 2019

Bibliographical note

Publisher Copyright:
© 2019 Elsevier B.V.

Funding

We thank Dr. K. Keinan-Adamsky, Dr. H. Gottlieb and Dr. M. Afri for spectrometer assistance, I. Tabakman for technical assistance, and Dr. I. Sher for her support in preparing LPNs. The MSP1D1-encoding plasmid was generously provided by Prof. Franz Hagn (Technische Universitat München). Establishment of the 700 MHz spectrometer system was supported by Fundácion Adar and a Converging Technologies award (Israel Science Foundation). GD and RZ acknowledge support by European grant PIRG4-GA-2008-239182 (T3SSBteA). JHC acknowledges support of the Christians for Israel Chair for Medical Research. We thank Dr. K. Keinan-Adamsky, Dr. H. Gottlieb and Dr. M. Afri for spectrometer assistance, I. Tabakman for technical assistance, and Dr. I. Sher for her support in preparing LPNs. The MSP1D1-encoding plasmid was generously provided by Prof. Franz Hagn (Technische Universitat M?nchen). Establishment of the 700?MHz spectrometer system was supported by Fund?cion Adar and a Converging Technologies award (Israel Science Foundation). GD and RZ acknowledge support by European grant PIRG4-GA-2008-239182 (T3SSBteA). JHC acknowledges support of the Christians for Israel Chair for Medical Research.

Funders	Funder number
Technische Universitat M?nchen
Israel Science Foundation	PIRG4-GA-2008-239182

Keywords

Bordetella
Effector
Membrane targeting
NMR spectroscopy
Type 3 secretion system
X-ray crystallography

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title = "Structure and membrane-targeting of a Bordetella pertussis effector N-terminal domain",

abstract = "BteA, a 69-kDa cytotoxic protein, is a type III secretion system (T3SS) effector in the classical Bordetella, the etiological agents of pertussis and related mammalian respiratory diseases. Like other cytotoxicity-mediating effectors, BteA uses its multifunctional N-terminal domain to target phosphatidylinositol (PI)-rich microdomains in the host membrane. Despite their structural similarity, T3SS effectors exhibit a variable range of membrane interaction modes, and currently only limited structural information is available for the BteA membrane-targeting domain and the molecular mechanisms underlying its function. Employing a synergistic combination of structural methods, here we determine the structure of this functional domain and uncover key molecular determinants mediating its interaction with membranes. Residues 29–121 of BteA form an elongated four-helix bundle packed against two shorter perpendicular helices, the second of which caps the domain in a critical {\textquoteleft}tip motif{\textquoteright}. A flexible region preceding the BteA helical bundle contains the characteristic β-motif required for binding its cognate chaperone BtcA. We show that BteA targets PI(4,5)P2-containing lipoprotein nanodiscs and binds a soluble PI(4,5)P2 analog via an extensive positively charged surface spanning its first two helices, and that this interaction is weaker for PI(3,5)P2 and abolished for PI(4)P. We confirmed this model of membrane-targeting by observation of BteA-induced changes in the structure of PI(4,5)P2-containing phospholipid bilayers using small-angle X-ray scattering (SAXS). We also extended these results to a larger BteA domain (residues 1–287), confirming its interaction with bilayers using calorimetry, fluorescence and SAXS methods. This novel view of the structural underpinnings of membrane targeting by BteA is an important step towards a comprehensive understanding of cytotoxicity in Bordetella, as well as interactions of a broad range of pathogens with their respective hosts.",

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author = "Adi Yahalom and Geula Davidov and Sofiya Kolusheva and Hadassa Shaked and Shiran Barber-Zucker and Raz Zarivach and Chill, {Jordan H.}",

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AU - Yahalom, Adi

AU - Davidov, Geula

AU - Kolusheva, Sofiya

AU - Shaked, Hadassa

AU - Barber-Zucker, Shiran

AU - Zarivach, Raz

AU - Chill, Jordan H.

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AB - BteA, a 69-kDa cytotoxic protein, is a type III secretion system (T3SS) effector in the classical Bordetella, the etiological agents of pertussis and related mammalian respiratory diseases. Like other cytotoxicity-mediating effectors, BteA uses its multifunctional N-terminal domain to target phosphatidylinositol (PI)-rich microdomains in the host membrane. Despite their structural similarity, T3SS effectors exhibit a variable range of membrane interaction modes, and currently only limited structural information is available for the BteA membrane-targeting domain and the molecular mechanisms underlying its function. Employing a synergistic combination of structural methods, here we determine the structure of this functional domain and uncover key molecular determinants mediating its interaction with membranes. Residues 29–121 of BteA form an elongated four-helix bundle packed against two shorter perpendicular helices, the second of which caps the domain in a critical ‘tip motif’. A flexible region preceding the BteA helical bundle contains the characteristic β-motif required for binding its cognate chaperone BtcA. We show that BteA targets PI(4,5)P2-containing lipoprotein nanodiscs and binds a soluble PI(4,5)P2 analog via an extensive positively charged surface spanning its first two helices, and that this interaction is weaker for PI(3,5)P2 and abolished for PI(4)P. We confirmed this model of membrane-targeting by observation of BteA-induced changes in the structure of PI(4,5)P2-containing phospholipid bilayers using small-angle X-ray scattering (SAXS). We also extended these results to a larger BteA domain (residues 1–287), confirming its interaction with bilayers using calorimetry, fluorescence and SAXS methods. This novel view of the structural underpinnings of membrane targeting by BteA is an important step towards a comprehensive understanding of cytotoxicity in Bordetella, as well as interactions of a broad range of pathogens with their respective hosts.

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Structure and membrane-targeting of a Bordetella pertussis effector N-terminal domain

Abstract

Bibliographical note

Funding

Keywords

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