Substrate-dependent transmembrane signaling in TonB-dependent transporters is not conserved

Kim, Mi‐Yeon; Fanucci, Gail E.; Cafiso, David S.

doi:10.1073/pnas.0702172104

Cited by 35 publications

(56 citation statements)

References 42 publications

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“…The TonB box serves as an energy-coupling motif. Electron paramagnetic resonance spectroscopy of the spin-labeled TonB box of isolated FhuA reveals that the TonB box is highly flexible, regardless of whether FhuA is substrate loaded or not (43). This stands in contrast to the TonB boxes of other transport-ers, which undergo substrate-induced order-to-disorder transitions (43).…”

Section: Energy-coupled Transport and Receptor Functions Of Fhuamentioning

confidence: 85%

FhuA (TonA), the Career of a Protein

Braun

2009

J Bacteriol

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The interest in the TonA protein, now called FhuA, has endured since the beginnings of phage genetics and molecular biology. The functions of this energy-coupled transporter and receptor in the outer membrane of Escherichia coli cells are fascinating. The historical perspective of this protein presented here is not intended to provide a comprehensive account of the structure and function of FhuA but rather attempts to explain how this protein has attracted interest for so long.The seminal work published by Salvador E. Luria and Max Delbrück (51) in 1943 is considered the beginning of modern bacterial genetics (12). In this publication, they reported their fluctuation test, which demonstrates that in bacteria, genetic mutations arise in the absence of selection rather than as a response to selection. They isolated virus-resistant mutants of Escherichia coli B, ascribed this virus resistance to mutations in the host cells prior to infection by the phage, and presented a means for quantitatively calculating mutation rates. The virus they used was later named phage T1, and the mutations that conferred resistance were subsequently designated tonA and tonB (ton derived from T one). It was later found that a phage morphologically and serologically distinct from T1, named T5, cannot infect tonA mutants but can infect tonB mutants; i.e., phage T5 requires only the tonA-encoded function but not the tonB-encoded function.Members of the "phage group" surrounding Luria and Delbrück were interested in genetics, reproduction, and multiplication. They studied, as the most simple system, phages and E. coli host cells, mutants, phage morphology, serology, adsorption, lysis, and burst size, but not biochemistry (12,14). The prejudice against biochemistry began to dissolve during the postdoctoral studies of Wolfhard Weidel, a biochemist by training, on phage adsorption and infection in Delbrück's laboratory. Delbrück's enthusiasm for Weidel's studies was evident in his report to the Caltech president, in which he referred to Weidel's work as "the most startling finding of the year" (66). Weidel's findings were included in the 1950 report on viruses of the Division of Biology of the California Institute of Technology. They were published in 1951 (68) after he was back in Germany at the Max Planck Institute of Biology in Tübingen, where he continued his work on phage adsorption, first with phages T2, T4, and T6 and then mainly with phage T5. His first achievement was the isolation of membrane fractions from E. coli B that bound the T phages with kinetics resembling those obtained with living bacteria. He could differentiate between receptors for T2, T4, and T6 on the one side and phage T5 on the other side. Electron microscopy revealed phage-induced degradation of the cell envelopes (68), which was later assigned to the T2 lysozyme (69).The methods used at that time were rather crude. The T5 receptor was extracted from cells by treatment with 0.1 N NaOH, neutralized with CO 2 , precipitated with 20% acetic acid, and then solubilized ...

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Section: Energy-coupled Transport and Receptor Functions Of Fhuamentioning

confidence: 85%

FhuA (TonA), the Career of a Protein

Braun

2009

J Bacteriol

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“…18 The Ton box conformation in FecA undergoes a substrate-dependent disordering that is sensitive to osmolytes Previous work using SDSL demonstrated that the Ton box of FecA, the Escherichia coli ferric citrate transporter, also displayed a substrate-dependent disorder transition on substrate addition. 28 Spectra from two sites in the Ton box of FecA, L82R1 and V85R1, are shown in Figure 7. Substrate addition narrows the lineshapes and increases the On the addition of PEG 1000 (following the addition of ferric citrate), the motion of R1 decreases dramatically, yielding spectra similar to but not identical to those of the substrate-free state [ Fig.…”

Section: Some Conformational Transitions In Btub Are Not Sensitive Tomentioning

confidence: 99%

Osmolytes modulate conformational exchange in solvent‐exposed regions of membrane proteins

Jiménez¹,

Cao²,

Kim³

et al. 2010

Protein Science

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Site-directed spin labeling (SDSL) was used to investigate local structure and conformational exchange in two bacterial outer-membrane TonB-dependent transporters, BtuB and FecA. Protecting osmolytes, such as polyethylene glycols (PEGs) are known to modulate a substrate-dependent conformational equilibrium in the energy coupling motif (Ton box) of BtuB. Here, we demonstrate that a segment that is N-terminal to the Ton box in BtuB, is in conformational exchange between ordered and disordered states with or without substrate. Protecting osmolytes shift this equilibrium to favor the more ordered, folded state. However, a segment of BtuB that is C-terminal to the Ton box that is not solvent exposed is insensitive to PEGs. Protecting osmolytes also modulate a conformational equilibrium in the Ton box of FecA, with larger molecular weight PEGs producing the largest shifts in the conformational free energy. These data indicate that solvent-exposed regions of these transporters undergo conformational exchange and that regions of these transporters that are involved in protein-protein interactions sample multiple conformational substates. The sensitivity to solute provides an explanation for differences seen between two high-resolution structures of BtuB, which each likely represent one conformation from a subset of states that are normally sampled by the protein. This work also illustrates how SDSL and osmolytes may be used to characterize and quantitate conformational equilibria in membrane proteins.

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“…Over the past three decades, many aspects of this TonB-ExbB-ExbD-dependent transport system have been revealed. The crystal structures of several OM transporters and their complexes with TonB are now known (Ferguson et al, 1998(Ferguson et al, , 2002Buchanan et al, 1999;Ferguson and Deisenhofer, 2004;Pawelek et al, 2006;Shultis et al, 2006;Krieg et al, 2009), the signal transduction of OM transporters by interaction with TonB has been elucidated (Ferguson et al, 2007;Kim et al, 2007) and the rotational mechanism of TonB motion has been reported (Jordan et al, 2013). However, with regard to the substrates of the transport system, we are probably only seeing the 'tip of the iceberg' (Schauer et al, 2008).…”

Section: Introductionmentioning

confidence: 99%

New insights into iron acquisition by cyanobacteria: an essential role for ExbB-ExbD complex in inorganic iron uptake

Jiang

Lou

et al. 2014

The ISME Journal

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Cyanobacteria are globally important primary producers that have an exceptionally large iron requirement for photosynthesis. In many aquatic ecosystems, the levels of dissolved iron are so low and some of the chemical species so unreactive that growth of cyanobacteria is impaired. Pathways of iron uptake through cyanobacterial membranes are now being elucidated, but the molecular details are still largely unknown. Here we report that the non-siderophore-producing cyanobacterium Synechocystis sp. PCC 6803 contains three exbB-exbD gene clusters that are obligatorily required for growth and are involved in iron acquisition. The three exbB-exbDs are redundant, but single and double mutants have reduced rates of iron uptake compared with wild-type cells, and the triple mutant appeared to be lethal. Short-term measurements in chemically well-defined medium show that iron uptake by Synechocystis depends on inorganic iron (Fe 0 ) concentration and ExbB-ExbD complexes are essentially required for the Fe 0 transport process. Although transport of iron bound to a model siderophore, ferrioxamine B, is also reduced in the exbB-exbD mutants, the rate of uptake at similar total [Fe] is about 800-fold slower than Fe 0 , suggesting that hydroxamate siderophore iron uptake may be less ecologically relevant than free iron. These results provide the first evidence that ExbB-ExbD is involved in inorganic iron uptake and is an essential part of the iron acquisition pathway in cyanobacteria. The involvement of an ExbB-ExbD system for inorganic iron uptake may allow cyanobacteria to more tightly maintain iron homeostasis, particularly in variable environments where iron concentrations range from limiting to sufficient.

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Substrate-dependent transmembrane signaling in TonB-dependent transporters is not conserved

Cited by 35 publications

References 42 publications

FhuA (TonA), the Career of a Protein

FhuA (TonA), the Career of a Protein

Osmolytes modulate conformational exchange in solvent‐exposed regions of membrane proteins

New insights into iron acquisition by cyanobacteria: an essential role for ExbB-ExbD complex in inorganic iron uptake

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