Estimating the size of the active translocation pore of an autotransporter.

A. Sauri, C.M. ten Hagen-Jongman, J.P. van Ulsen, S. Luirink

Research output: Contribution to JournalArticleAcademic

Abstract

Autotransporters (ATs) are large virulence factors secreted by Gram-negative bacteria. The passenger domain, carrying the virulence functions, is transported across the bacterial outer membrane in a step that is facilitated by a C-terminal β-domain. This domain folds into a β-barrel with a central aqueous pore of ∼ 1 nm inner diameter according to crystal structures. However, these static dimensions are not compatible with the observed secretion of passengers that may contain natural short-spaced disulfide bonds or artificially fused folded elements. Here, we have systematically analyzed the dimensions of the active AT passenger translocator by inserting peptides of different length and structural complexity in the passenger of the AT hemoglobin protease. The peptides were introduced in a short loop protruding from the main structure and flanked by two single cysteines. Our results show that the attained secondary structure may be more critical for secretion than the length of peptide inserted. Furthermore, the data suggest that, during passenger translocation, at least four extended polypeptides or an extended polypeptide and an α-helix are accommodated in the translocator, indicating that the diameter of the active translocation pore is up to 1.7 nm. If the β-domain functions as the translocator, it must be forced into an expanded conformation during passenger translocation. © 2011 Elsevier Ltd. All rights reserved.
Original languageEnglish
Pages (from-to)335-345
JournalJournal of Molecular Biology
Volume416
Issue number3
Early online date29 Dec 2011
DOIs
Publication statusPublished - 2012

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Peptides
Virulence Factors
Gram-Negative Bacteria
Disulfides
Cysteine
Virulence
Hemoglobins
Peptide Hydrolases
Type V Secretion Systems
Membranes

Cite this

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title = "Estimating the size of the active translocation pore of an autotransporter.",
abstract = "Autotransporters (ATs) are large virulence factors secreted by Gram-negative bacteria. The passenger domain, carrying the virulence functions, is transported across the bacterial outer membrane in a step that is facilitated by a C-terminal β-domain. This domain folds into a β-barrel with a central aqueous pore of ∼ 1 nm inner diameter according to crystal structures. However, these static dimensions are not compatible with the observed secretion of passengers that may contain natural short-spaced disulfide bonds or artificially fused folded elements. Here, we have systematically analyzed the dimensions of the active AT passenger translocator by inserting peptides of different length and structural complexity in the passenger of the AT hemoglobin protease. The peptides were introduced in a short loop protruding from the main structure and flanked by two single cysteines. Our results show that the attained secondary structure may be more critical for secretion than the length of peptide inserted. Furthermore, the data suggest that, during passenger translocation, at least four extended polypeptides or an extended polypeptide and an α-helix are accommodated in the translocator, indicating that the diameter of the active translocation pore is up to 1.7 nm. If the β-domain functions as the translocator, it must be forced into an expanded conformation during passenger translocation. {\circledC} 2011 Elsevier Ltd. All rights reserved.",
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Estimating the size of the active translocation pore of an autotransporter. / Sauri, A.; ten Hagen-Jongman, C.M.; van Ulsen, J.P.; Luirink, S.

In: Journal of Molecular Biology, Vol. 416, No. 3, 2012, p. 335-345.

Research output: Contribution to JournalArticleAcademic

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AU - van Ulsen, J.P.

AU - Luirink, S.

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N2 - Autotransporters (ATs) are large virulence factors secreted by Gram-negative bacteria. The passenger domain, carrying the virulence functions, is transported across the bacterial outer membrane in a step that is facilitated by a C-terminal β-domain. This domain folds into a β-barrel with a central aqueous pore of ∼ 1 nm inner diameter according to crystal structures. However, these static dimensions are not compatible with the observed secretion of passengers that may contain natural short-spaced disulfide bonds or artificially fused folded elements. Here, we have systematically analyzed the dimensions of the active AT passenger translocator by inserting peptides of different length and structural complexity in the passenger of the AT hemoglobin protease. The peptides were introduced in a short loop protruding from the main structure and flanked by two single cysteines. Our results show that the attained secondary structure may be more critical for secretion than the length of peptide inserted. Furthermore, the data suggest that, during passenger translocation, at least four extended polypeptides or an extended polypeptide and an α-helix are accommodated in the translocator, indicating that the diameter of the active translocation pore is up to 1.7 nm. If the β-domain functions as the translocator, it must be forced into an expanded conformation during passenger translocation. © 2011 Elsevier Ltd. All rights reserved.

AB - Autotransporters (ATs) are large virulence factors secreted by Gram-negative bacteria. The passenger domain, carrying the virulence functions, is transported across the bacterial outer membrane in a step that is facilitated by a C-terminal β-domain. This domain folds into a β-barrel with a central aqueous pore of ∼ 1 nm inner diameter according to crystal structures. However, these static dimensions are not compatible with the observed secretion of passengers that may contain natural short-spaced disulfide bonds or artificially fused folded elements. Here, we have systematically analyzed the dimensions of the active AT passenger translocator by inserting peptides of different length and structural complexity in the passenger of the AT hemoglobin protease. The peptides were introduced in a short loop protruding from the main structure and flanked by two single cysteines. Our results show that the attained secondary structure may be more critical for secretion than the length of peptide inserted. Furthermore, the data suggest that, during passenger translocation, at least four extended polypeptides or an extended polypeptide and an α-helix are accommodated in the translocator, indicating that the diameter of the active translocation pore is up to 1.7 nm. If the β-domain functions as the translocator, it must be forced into an expanded conformation during passenger translocation. © 2011 Elsevier Ltd. All rights reserved.

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