Characterization of TonB Interactions with the FepA Cork Domain and FecA N-terminal Signaling Domain |
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Authors: | R Sean Peacock Valery V Andrushchenko A Ross Demcoe Matt Gehmlich Lily Sia Lu Alicia Garcia Herrero Hans J Vogel |
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Institution: | (1) Structural Biology Research Group, Department of Biological Sciences, University of Calgary, Calgary, Alberta, T2N 1N4, Canada |
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Abstract: | The mechanism of TonB dependent siderophore uptake through outer membrane transporters in Gram-negative bacteria is poorly
understood. In an effort to expand our knowledge of the interaction between TonB and the outer membrane transporters, we have
cloned and expressed the FepA cork domain (11–154) from Salmonella typhimurium and characterized its interaction with the periplasmic C-terminal domain of TonB (103–239) by isotope assisted FTIR and NMR
spectroscopy. For comparison we also performed similar experiments using the FecA N-terminal domain (1–96) from Escherichia coli which includes the conserved TonB box. The FepA cork domain was completely unfolded in solution, as observed for the E. coli cork domain previously Usher et al. (2001) Proc Natl Acad Sci USA 98, 10676–10681]. The FepA cork domain was found to bind to TonB, eliciting essentially the same chemical shift changes in TonB
C-terminal domain as was observed in the presence of TonB box peptides. The FecA construct did not cause this same structural
change in TonB. The binding of the FepA cork domain to TonB-CTD was found to decrease the amount of ordered secondary structure
in TonB-CTD. It is likely that the FecA N-terminal domain interferes with TonB-CTD binding to the TonB box. Binding of the
FepA cork domain induces a loss of secondary structure in TonB, possibly exposing TonB surface area for additional intermolecular
interactions such as potential homodimerization or additional interactions with the barrel of the outer membrane transporter. |
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Keywords: | fourier transfrom infrared spectroscopy Gram-negative bacteria iron transport nuclear magnetic resonance TonB |
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