CryoEM and computer simulations reveal a novel kinase conformational switch in bacterial chemotaxis signaling

Cassidy, C. Keith; Himes, Benjamin A.; Alvarez, Frances Joan D.; Ma, Jun; Zhao, Gongpu; Perilla, Juan R.; Schulten, Klaus; Zhang, Peijun

doi:10.7554/elife.08419

Cited by 106 publications

(244 citation statements)

References 62 publications

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“…Cryo-electron tomography revealed the existence of CheW rings symbolic of CheW-CheW interactions in E. coli chemoreceptor array structures (37). We suggest a similar chemotaxis array in H. pylori in which CheW-CheV1 rings form the core of the complex.…”

Section: Discussionmentioning

confidence: 78%

Cooperation of two distinct coupling proteins creates chemosensory network connections

Abedrabbo

Castellon

Collins

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Although it is appreciated that bacterial chemotaxis systems rely on coupling, also called scaffold, proteins to both connect input receptors with output kinases and build interkinase connections that allow signal amplification, it is not yet clear why many systems use more than one coupling protein. We examined the distinct functions for multiple coupling proteins in the bacterial chemotaxis system of Helicobacter pylori, which requires two nonredundant coupling proteins for chemotaxis: CheW and CheV1, a hybrid of a CheW and a phosphorylatable receiver domain. We report that CheV1 and CheW have largely redundant abilities to interact with chemoreceptors and the CheA kinase, and both similarly activated CheA’s kinase activity. We discovered, however, that they are not redundant for formation of the higher order chemoreceptor arrays that are known to form via CheA–CheW interactions. In support of this possibility, we found that CheW and CheV1 interact with each other and with CheA independent of the chemoreceptors. Therefore, it seems that some microbes have modified array formation to require CheW and CheV1. Our data suggest that multiple coupling proteins may be used to provide flexibility in the chemoreceptor array formation.

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Section: Discussionmentioning

confidence: 78%

Cooperation of two distinct coupling proteins creates chemosensory network connections

Abedrabbo

Castellon

Collins

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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“…For this assay, our "wild type" CheA protein carried serine replacements at the three native, but nonessential, CheA cysteine residues (16), along with HA affinity tags in the P1-P2 and P2-P3 linkers (17,18), to detect CheA cross-linking products in anti-HA immunoblots. The most efficient and specific interface 2 reporter (7,8,10). In addition to P5/CheW rings (dashed black lines), six-member CheW rings interact with the receptor dimers that do not contact CheA or CheW in the component core complexes (7,10).…”

Section: Resultsmentioning

confidence: 99%

“…The most efficient and specific interface 2 reporter (7,8,10). In addition to P5/CheW rings (dashed black lines), six-member CheW rings interact with the receptor dimers that do not contact CheA or CheW in the component core complexes (7,10). CheA-P5•CheW interface 2 was based on crystal structures of T. maritima proteins (9).…”

Section: Resultsmentioning

confidence: 99%

“…The architecture of the chemosensory array has been elucidated at the molecular level by fitting the atomic coordinates of individual proteins into electron density maps obtained by cryoelectron tomography (7)(8)(9)(10). These studies, in conjunction with targeted crosslinking to confirm protein-protein interfaces (11,12), generated a structural model for the core signaling complex and its higher-order organization in the array (Fig.…”

mentioning

confidence: 99%

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The source of high signal cooperativity in bacterial chemosensory arrays

Piñas

Frank

Vaknin

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

127

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The Escherichia coli chemosensory system consists of large arrays of transmembrane chemoreceptors associated with a dedicated histidine kinase, CheA, and a linker protein, CheW, that couples CheA activity to receptor control. The kinase activity responses to receptor ligand occupancy changes can be highly cooperative, reflecting allosteric coupling of multiple CheA and receptor molecules. Recent structural and functional studies have led to a working model in which receptor core complexes, the minimal units of signaling, are linked into hexagonal arrays through a unique interface 2 interaction between CheW and the P5 domain of CheA. To test this array model, we constructed and characterized CheA and CheW mutants with amino acid replacements at key interface 2 residues. The mutant proteins proved defective in interface 2-specific in vivo cross-linking assays, and formed signaling complexes that were dispersed around the cell membrane rather than clustered at the cell poles as in wild type chemosensory arrays. Interface 2 mutants down-regulated CheA activity in response to attractant stimuli in vivo, but with much less cooperativity than the wild type. Moreover, mutant cells containing fluorophore-tagged receptors exhibited greater basal anisotropy that changed rapidly in response to attractant stimuli, consistent with facile changes in loosely packed receptors. We conclude that interface 2 lesions disrupt important network connections between core complexes, preventing receptors from operating in large, allosteric teams. This work confirms the critical role of interface 2 in organizing the chemosensory array, in directing the clustered array to the cell poles, and in producing its highly cooperative signaling properties.chemotaxis | chemoreceptors | kinase activity | stimulus response

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“…The combination of such structural data with molecular dynamics (MD) simulation has been shown to provide a powerful approach by which to gain insight into the function of molecular machines (72). Briefly, MD simulation is a computational method that uses an empirically based potential energy function to Recently, a combination of experimental and computational techniques has been applied to chemotaxis arrays (73). Here, crystallographic structures of an individual receptor, CheA, and CheW proteins from Thermotoga maritima were assembled and refined according to cryo-EM maps of the extended E. coli signaling complex by MD flexible fitting (74), a technique based on MD simulation.…”

Section: Expanding the Toolboxmentioning

confidence: 99%

Recent Advances and Future Prospects in Bacterial and Archaeal Locomotion and Signal Transduction

et al. 2017

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The structure and function of two-component and chemotactic signaling and different aspects related to the motility of bacteria and archaea are key research areas in modern microbiology. Escherichia coli is the traditional model organism used to study chemotaxis signaling and motility. However, the recent study of a wide range of bacteria and even some archaea with different lifestyles has provided new insight into the ecophysiology of chemotaxis, which is essential for the establishment of different pathogens or beneficial bacteria in a host. The expanded range of model organisms has also permitted the study of chemosensory pathways unrelated to chemotaxis, multiple chemotaxis pathways within an organism, and new types of chemoreceptors. This research has greatly benefitted from technical advances in the field of cryomicroscopy, which continues to reveal with increasing resolution the complexity and diversity of large protein complexes like the flagellar motor or chemoreceptor arrays. In addition, sensitive instruments now allow an increasing number of experiments to be conducted at the single-cell level, thereby revealing information that is beginning to bridge the gap between individual cells and population behavior. Evidence has also accumulated showing that bacteria have evolved different mechanisms for surface sensing, which appears to be mediated by flagella and possibly type IV pili, and that the downstream signaling involves chemosensory pathways and two-component-system-based processes. Herein, we summarize the recent advances and research tendencies in this field as presented at the latest Bacterial Locomotion and Signal Transduction (BLAST XIV) conference.KEYWORDS chemotaxis, flagella, flagellar motility, signal transduction, two-component regulatory systems T he capacity to sense and respond to changes in environmental cues is an essential feature of the prokaryotic lifestyle. As a consequence, bacteria and archaea have evolved an array of different molecular mechanisms that permit the detection of signals in order to generate appropriate cellular responses. These responses are mediated primarily by one-and two-component systems, as well as chemosensory signaling pathways (1-4). Whereas the former systems mediate changes primarily at the transcriptional level, chemosensory pathways form the basis for chemotaxis, the directed movement of prokaryotes in compound gradients. The study of signaling processes is not only of fundamental interest but may also contribute to the tackling of one of the central clinical problems, which is the increasing amount of antibiotic-resistant pathogens. There is now a significant amount of data indicating that interference with signal transduction systems, motility, and chemotaxis can be an alternative strategy to weaken or block pathogens (5, 6).In

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CryoEM and computer simulations reveal a novel kinase conformational switch in bacterial chemotaxis signaling

Cited by 106 publications

References 62 publications

Cooperation of two distinct coupling proteins creates chemosensory network connections

Cooperation of two distinct coupling proteins creates chemosensory network connections

The source of high signal cooperativity in bacterial chemosensory arrays

Recent Advances and Future Prospects in Bacterial and Archaeal Locomotion and Signal Transduction

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