Cross-Linking Evidence for Motional Constraints within Chemoreceptor Trimers of Dimers

Massazza, Diego A.; Parkinson, John S.; Studdert, Claudia A.

doi:10.1021/bi101483r

Cited by 13 publications

(14 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…20,25,29 Using in vitro conditions with vesicle template assembly offers a complementary look at this crosslinking reaction. Initial experiments with this crosslinking agent examined the concentration dependence of dimer and trimer formation to choose optimal concentrations for further experiments.…”

Section: Resultsmentioning

confidence: 99%

Membrane Association of a Protein Increases the Rate, Extent, and Specificity of Chemical Cross-Linking

et al. 2013

View full text Add to dashboard Cite

Many cellular processes involve interactions between membrane-associated proteins, and those interactions are enhanced by membrane association. We have used crosslinking reactions to compare the extent and specificity of protein interactions in solution versus on a membrane surface. Cysteine mutants of a soluble cytoplasmic fragment (CF) of the aspartate receptor, a transmembrane receptor involved in bacterial chemotaxis, are used in disulfide bond formation with the thiol-specific oxidant diamide and chemical crosslinking reactions with the trifunctional maleimide TMEA. CF binding to vesicles is mediated by its N-terminal His tag binding to vesicles containing a nickel-chelating lipid, so crosslinking reactions conducted in the presence and absence of vesicles differ only in whether CF is bound to the vesicles or is free in solution. For multiple Cys throughout the CF, membrane association is shown to increase the rate and extent of these reactions. Crosslinking specificity, which is measured as the preference for crosslinking between Cys near each other in the native structure, is also enhanced by membrane association. These results provide an experimental demonstration that membrane binding enhances protein-protein interactions, an important consideration for understanding processes involving membrane-associated proteins. The experiments further demonstrate the importance of crosslinking conditions for these reactions that are often used to probe protein structure and dynamics, and the potential of membrane association to restore native interactions of membrane-associated proteins for crosslinking studies.

show abstract

Section: Resultsmentioning

confidence: 99%

Membrane Association of a Protein Increases the Rate, Extent, and Specificity of Chemical Cross-Linking

et al. 2013

View full text Add to dashboard Cite

show abstract

“…The c-di-GMP-induced conformational changes may affect Tlp1 interactions with the downstream chemotaxis signaling complex and/or with other chemotaxis receptors that are known to form mixed multireceptor arrays (27–32). …”

Section: Discussionmentioning

confidence: 99%

Integration of the Second Messenger c-di-GMP into the Chemotactic Signaling Pathway

Russell

Bible

Fang

et al. 2013

mBio

View full text Add to dashboard Cite

Elevated intracellular levels of the bacterial second messenger c-di-GMP are known to suppress motility and promote sessility. Bacterial chemotaxis guides motile cells in gradients of attractants and repellents over broad concentration ranges, thus allowing bacteria to quickly adapt to changes in their surroundings. Here, we describe a chemotaxis receptor that enhances, as opposed to suppresses, motility in response to temporary increases in intracellular c-di-GMP. Azospirillum brasilense’s preferred metabolism is adapted to microaerophily, and these motile cells quickly navigate to zones of low oxygen concentration by aerotaxis. We observed that changes in oxygen concentration result in rapid changes in intracellular c-di-GMP levels. The aerotaxis and chemotaxis receptor, Tlp1, binds c-di-GMP via its C-terminal PilZ domain and promotes persistent motility by increasing swimming velocity and decreasing swimming reversal frequency, which helps A. brasilense reach low-oxygen zones. If c-di-GMP levels remain high for extended periods, A. brasilense forms nonmotile clumps or biofilms on abiotic surfaces. These results suggest that association of increased c-di-GMP levels with sessility is correct on a long-term scale, while in the short-term c-di-GMP may actually promote, as opposed to suppress, motility. Our data suggest that sensing c-di-GMP by Tlp1 functions similar to methylation-based adaptation. Numerous chemotaxis receptors contain C-terminal PilZ domains or other sensory domains, suggesting that intracellular c-di-GMP as well as additional stimuli can be used to modulate adaptation of bacterial chemotaxis receptors.

show abstract

“…Although chemoreceptors do not have any obvious structural similarity to well-known membrane curvature sensing and generating domains16, electron microscopy has shown that overexpressed E. coli serine chemoreceptor, Tsr, forms tubular membrane invaginations and apparent vesicular structures17. Subsequent cryo-electron tomography1819 and cross-linking20 have found that chemoreceptors form a cone-shaped trimer-of-dimers (ToD) in vivo , corroborating a previous crystal structure of the cytoplasmic domain of Tsr21. This ToD geometry led Endres22 to propose a curvature-based mechanism for polar localization of the array.…”

mentioning

confidence: 99%

Origins of chemoreceptor curvature sorting in Escherichia coli

Draper

Liphardt

2017

Nat Commun

View full text Add to dashboard Cite

Bacterial chemoreceptors organize into large clusters at the cell poles. Despite a wealth of structural and biochemical information on the system's components, it is not clear how chemoreceptor clusters are reliably targeted to the cell pole. Here, we quantify the curvature-dependent localization of chemoreceptors in live cells by artificially deforming growing cells of Escherichia coli in curved agar microchambers, and find that chemoreceptor cluster localization is highly sensitive to membrane curvature. Through analysis of multiple mutants, we conclude that curvature sensitivity is intrinsic to chemoreceptor trimers-of-dimers, and results from conformational entropy within the trimer-of-dimers geometry. We use the principles of the conformational entropy model to engineer curvature sensitivity into a series of multi-component synthetic protein complexes. When expressed in E. coli, the synthetic complexes form large polar clusters, and a complex with inverted geometry avoids the cell poles. This demonstrates the successful rational design of both polar and anti-polar clustering, and provides a synthetic platform on which to build new systems.

show abstract

Cross-Linking Evidence for Motional Constraints within Chemoreceptor Trimers of Dimers

Cited by 13 publications

References 32 publications

Membrane Association of a Protein Increases the Rate, Extent, and Specificity of Chemical Cross-Linking

Membrane Association of a Protein Increases the Rate, Extent, and Specificity of Chemical Cross-Linking

Integration of the Second Messenger c-di-GMP into the Chemotactic Signaling Pathway

Origins of chemoreceptor curvature sorting in Escherichia coli

Contact Info

Product

Resources

About