Taking the Escherichia coli TonB transmembrane domain "Offline"? Nonprotonatable Asn substitutes fully for TonB his20

Cheryl Swayne, Kathleen Postle

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

The TonB system of Gram-negative bacteria uses the proton motive force (PMF) of the cytoplasmic membrane to energize active transport of nutrients across the outer membrane. The single transmembrane domain (TMD) anchor of TonB, the energy transducer, is essential. Within that TMD, His20 is the only TMD residue that is unable to withstand alanine replacement without a loss of activity. H20 is required for a PMF-dependent conformational change, suggesting that the importance of H20 lies in its ability to be reversibly protonated and deprotonated. Here all possible residues were substituted at position 20 (H20X substitutions). The His residue was also relocated throughout the TonB TMD. Surprisingly, Asn, a structurally similar but nonprotonatable residue, supported full activity at position 20; H20S was very weakly active. All the remaining substitutions, including H20K, H20R, H20E, and H20D, the obvious candidates to mimic a protonated state or support proton translocation, were inactive. A second-site suppressor, ExbB(A39E), indiscriminately reactivated the majority of H20 substitutions and relocations, including H20V, which cannot be made protonatable. These results suggested that the TonB TMD was not on a proton conductance pathway and thus only indirectly responds to PMF, probably via ExbD.

Original languageEnglish (US)
Pages (from-to)3693-3701
Number of pages9
JournalJournal of bacteriology
Volume193
Issue number15
DOIs
StatePublished - Aug 1 2011

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Proton-Motive Force
Escherichia coli
Protons
Active Biological Transport
Gram-Negative Bacteria
Transducers
Alanine
Cell Membrane
Food
Membranes

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Biology

Cite this

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abstract = "The TonB system of Gram-negative bacteria uses the proton motive force (PMF) of the cytoplasmic membrane to energize active transport of nutrients across the outer membrane. The single transmembrane domain (TMD) anchor of TonB, the energy transducer, is essential. Within that TMD, His20 is the only TMD residue that is unable to withstand alanine replacement without a loss of activity. H20 is required for a PMF-dependent conformational change, suggesting that the importance of H20 lies in its ability to be reversibly protonated and deprotonated. Here all possible residues were substituted at position 20 (H20X substitutions). The His residue was also relocated throughout the TonB TMD. Surprisingly, Asn, a structurally similar but nonprotonatable residue, supported full activity at position 20; H20S was very weakly active. All the remaining substitutions, including H20K, H20R, H20E, and H20D, the obvious candidates to mimic a protonated state or support proton translocation, were inactive. A second-site suppressor, ExbB(A39E), indiscriminately reactivated the majority of H20 substitutions and relocations, including H20V, which cannot be made protonatable. These results suggested that the TonB TMD was not on a proton conductance pathway and thus only indirectly responds to PMF, probably via ExbD.",
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Taking the Escherichia coli TonB transmembrane domain "Offline"? Nonprotonatable Asn substitutes fully for TonB his20. / Swayne, Cheryl; Postle, Kathleen.

In: Journal of bacteriology, Vol. 193, No. 15, 01.08.2011, p. 3693-3701.

Research output: Contribution to journalArticle

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