scholarly journals Structural insight into the role of the PAS domainfor signal transduction in sensor-kinase BvgS

2021 ◽  
Author(s):  
Elian Dupré ◽  
Bernard Clantin ◽  
Youhua Yuan ◽  
Sophie Lecher ◽  
Elodie Lesne ◽  
...  
Keyword(s):  
Bordetella Pertussis ◽  
Whooping Cough ◽  
Sensory Transduction ◽  
Dimensional Structure ◽  
Sensor Kinase ◽  
Pas Domain ◽  
Human Respiratory Tract ◽  
Histidine Kinases ◽  
X Ray ◽  
X Ray Crystallography

The two-component system BvgAS controls the virulence regulon in Bordetella pertussis. BvgS is the prototype of a family of sensor histidine-kinases harboring periplasmic Venus flytrap (VFT) domains. The VFT domains are connected to the cytoplasmic kinase moiety by helical linkers separated by a Per-ARNT-Sim (PAS) domain. Antagonism between the two linkers, as one forms a coiled coil when the other is dynamic and vice versa, regulates BvgS activity. Here we solved the structure of the intervening PAS domain by X-ray crystallography. Two forms were obtained that notably differ by the connections between the PAS core domain and the flanking helical linkers. Structure-guided mutagenesis indicated that those connections participate in the regulation of BvgS activity. The PAS domain thus appears to function as a switch-facilitator module whose conformation determines the output of the system. As many BvgS homologs have similar architectures, the mechanisms unveiled here are likely to generally apply to the regulation of sensor-histidine kinases of that family. IMPORTANCE The whooping cough agent Bordetella pertussis colonizes the human respiratory tract using virulence factors co-regulated by the sensory transduction system BvgAS. BvgS is a model for a family of sensor-kinase proteins, some of which are found in important bacterial pathogens. BvgS functions as a kinase or a phosphatase depending on external signals, which determines if B. pertussis is virulent or avirulent. Deciphering its mode of action might thus lead to new ways of fighting infections. Here we used X-ray crystallography to solve the three-dimensional structure of the domain that precedes the enzymatic moiety and identified features that regulate BvgS activity. As many sensor-kinases of the BvgS family harbor homologous domains, the mechanism unveiled here might be of general relevance.

scholarly journals Role of Computational Methods in Going beyond X-ray Crystallography to Explore Protein Structure and Dynamics

2018 ◽  
Vol 19 (11) ◽  
pp. 3401 ◽  
Author(s):  
Ashutosh Srivastava ◽  
Tetsuro Nagai ◽  
Arpita Srivastava ◽  
Osamu Miyashita ◽  
Florence Tama
Keyword(s):  
Protein Dynamics ◽  
Computational Methods ◽  
Protein Structures ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
X Ray ◽  
X Ray Crystallography ◽  
Insight Into

Protein structural biology came a long way since the determination of the first three-dimensional structure of myoglobin about six decades ago. Across this period, X-ray crystallography was the most important experimental method for gaining atomic-resolution insight into protein structures. However, as the role of dynamics gained importance in the function of proteins, the limitations of X-ray crystallography in not being able to capture dynamics came to the forefront. Computational methods proved to be immensely successful in understanding protein dynamics in solution, and they continue to improve in terms of both the scale and the types of systems that can be studied. In this review, we briefly discuss the limitations of X-ray crystallography in studying protein dynamics, and then provide an overview of different computational methods that are instrumental in understanding the dynamics of proteins and biomacromolecular complexes.

scholarly journals Topography Prediction of Helical Transmembrane Proteins by a New Modification of the Sliding Window Method

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Maria N. Simakova ◽  
Nikolai N. Simakov
Keyword(s):  
Membrane Proteins ◽  
Transmembrane Domain ◽  
Domain Boundary ◽  
Three Dimensional ◽  
Transmembrane Proteins ◽  
Sliding Window ◽  
Dimensional Structure ◽  
X Ray ◽  
X Ray Crystallography ◽  
Window Method

Protein functions are specified by its three-dimensional structure, which is usually obtained by X-ray crystallography. Due to difficulty of handling membrane proteins experimentally to date the structure has only been determined for a very limited part of membrane proteins (<4%). Nevertheless, investigation of structure and functions of membrane proteins is important for medicine and pharmacology and, therefore, is of significant interest. Methods of computer modeling based on the data on the primary protein structure or the symbolic amino acid sequence have become an actual alternative to the experimental method of X-ray crystallography for investigating the structure of membrane proteins. Here we presented the results of the study of 35 transmembrane proteins, mainly GPCRs, using the novel method of cascade averaging of hydrophobicity function within the limits of a sliding window. The proposed method allowed revealing 139 transmembrane domains out of 140 (or 99.3%) identified by other methods. Also 236 transmembrane domain boundary positions out of 280 (or 84%) were predicted correctly by the proposed method with deviation from the predictions made by other methods that does not exceed the detection error of this method.

scholarly journals A new on-axis multimode spectrometer for the macromolecular crystallography beamlines of the Swiss Light Source

2009 ◽  
Vol 16 (2) ◽  
pp. 173-182 ◽  
Author(s):  
Robin L. Owen ◽  
Arwen R. Pearson ◽  
Alke Meents ◽  
Pirmin Boehler ◽  
Vincent Thominet ◽  
...  
Keyword(s):  
Light Source ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
X Ray ◽  
X Ray Crystallography ◽  
Additional Information ◽  
Swiss Light Source ◽  
Induced Changes ◽  
Insight Into

X-ray crystallography at third-generation synchrotron sources permits tremendous insight into the three-dimensional structure of macromolecules. Additional information is, however, often required to aid the transition from structure to function. In situ spectroscopic methods such as UV–Vis absorption and (resonance) Raman can provide this, and can also provide a means of detecting X-ray-induced changes. Here, preliminary results are introduced from an on-axis UV–Vis absorption and Raman multimode spectrometer currently being integrated into the beamline environment at X10SA of the Swiss Light Source. The continuing development of the spectrometer is also outlined.

scholarly journals Induction of Rare Conformation of Oligosaccharide by Binding to Calcium-dependent Bacterial Lectin: X-ray Crystallography and Modelling Study

2019 ◽  
Author(s):  
Martin Lepsik ◽  
Roman Sommer ◽  
Sakonwan Kuhaudomlarp ◽  
Mickaёl Lelimousin ◽  
Emanuele Paci ◽  
...  
Keyword(s):  
Force Field ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Lewis X ◽  
X Ray ◽  
X Ray Crystallography ◽  
Calcium Dependent ◽  
Protein Receptors ◽  
Dependent Protein ◽  
Micro Organisms

ABSTRACTPathogenic micro-organisms utilize protein receptors in adhesion to host tissues, a process that in some cases relies on the interaction between lectin and human glycoconjugates. Oligosaccharide epitopes are recognized through their three-dimensional structure and their flexibility is a key issue in specificity. In this paper, we analyse by X-ray crystallography the structures of the lectin LecB from two strains of Pseudomonas aeruginosa in complex with Lewis x oligosaccharide present on cell surfaces of human tissues. An unusual conformation of the glycan was observed in all binding sites with a non-canonical syn orientation of the N-acetyl group of N-acetyl-glucosamine. A PDB-wide search revealed that such an orientation occurs only in 2% of protein/carbohydrate complexes. Theoretical chemistry calculations showed that the observed conformation is unstable in solution but stabilised by the lectin. A reliable description of LecB/Lewis x complex by force field-based methods had proven as especially challenging due to the special feature of the binding site, two closely apposed Ca2+ ions which induce strong charge delocalisation. By comparing various force-field parametrisations, we design general protocols which will be useful in near future for designing carbohydrate-based ligands (glycodrugs) against other calcium-dependent protein receptors.

Synopses from the poster exhibition organized by I. M. Kerr, 1. Interferon: a tertiary structure predicted from amino acid sequences

1982 ◽  
Vol 299 (1094) ◽  
pp. 125-127 ◽  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Tertiary Structure ◽  
Three Dimensional ◽  
Amino Acid Sequences ◽  
Dimensional Structure ◽  
X Ray ◽  
Three Dimensional Structure ◽  
X Ray Crystallography ◽  
Functional Studies

Functional studies on interferon would be helped by a three-dimensional structure for the molecule. However, it may be several years before the structure of the protein is determined by X-ray crystallography. We have therefore used available methods for predicting the secondary - and the tertiary - structure of a protein from its amino acid sequence to propose a tertiary model involving the packing of four a-helices. Details of this work have been published elsewhere (Sternberg & Cohen 1982).

scholarly journals Coiled-Coil Antagonism Regulates Activity of Venus Flytrap-Domain-Containing Sensor Kinases of the BvgS Family

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Elodie Lesne ◽  
Elian Dupré ◽  
Marc F. Lensink ◽  
Camille Locht ◽  
Rudy Antoine ◽  
...  
Keyword(s):  
Bordetella Pertussis ◽  
Coiled Coil ◽  
Sensory Transduction ◽  
The Other ◽  
Pas Domain ◽  
Two Component System ◽  
Component System ◽  
Venus Flytrap ◽  
Two Component ◽  
Sensor Kinases

ABSTRACTBordetella pertussiscontrols the expression of its virulence regulon through the two-component system BvgAS. BvgS is a prototype for a family of multidomain sensor kinases. In BvgS, helical linkers connect periplasmic Venus flytrap (VFT) perception domains to a cytoplasmic Per-Arnt-Sim (PAS) domain and the PAS domain to the dimerization/histidine phosphotransfer (DHp) domain of the kinase. The two linkers can adopt coiled-coil structures but cannot do so simultaneously. The first linker forms a coiled coil in the kinase mode and the second in the phosphatase mode, with the other linker in both cases showing an increase in dynamic behavior. The intervening PAS domain changes its quaternary structure between the two modes. In BvgS homologues without a PAS domain, a helical “X” linker directly connects the VFT and DHp domains. Here, we used BvgS as a platform to characterize regulation in members of the PAS-less subfamily. BvgS chimeras of homologues with natural X linkers displayed various regulation phenotypes. We identified two distinct coiled-coil registers in the N- and C-terminal portions of the X linkers. Stable coil formation in the C-terminal moiety determines the phosphatase mode, similarly to BvgS; in contrast, coil formation in the N-terminal moiety along the other register leads to the kinase mode. Thus, antagonism between two registers in the VFT-DHp linker forms the basis for activity regulation in the absence of the PAS domain. The N and C moieties of the X linker play roles similar to those played by the two independent linkers in sensor kinases with a PAS domain, providing a unified mechanism of regulation for the entire family.IMPORTANCEThe whooping cough agentBordetella pertussisuses the BvgAS sensory transduction two-component system to regulate production of its virulence factors. BvgS serves as a model for a large family of multidomain bacterial sensor kinases.B. pertussisis virulent when BvgS functions as a kinase and avirulent when it switches to phosphatase activity in response to modulating signals. Understanding the molecular regulation of those proteins might lead to new antibacterial strategies. Here, we show that the linker regions between the perception and the enzymatic domains shift between distinct states of conformation in an alternating manner in response to signals and that their antagonistic changes control sensor kinase activity. These linker regions and mechanistic principles appear to be conserved among BvgS homologues, irrespective of the presence or absence of an intervening domain between the perception and the enzymatic domains. This work has thus uncovered general molecular mechanisms that regulate activity of sensor kinases in the BvgS family.

scholarly journals The BvgS PAS domain: an independent sensory perception module in theBordetellaBvgAS phosphorelay

2019 ◽  
Author(s):  
M. Ashley Sobran ◽  
Peggy A. Cotter
Keyword(s):  
Respiratory Tract ◽  
Bordetella Pertussis ◽  
Kinase Activity ◽  
Expression Patterns ◽  
Pas Domain ◽  
Human Respiratory Tract ◽  
Signal Perception ◽  
Two Component

AbstractTo detect and respond to the diverse environments they encounter, bacteria often use two-component regulatory systems (TCSs) to coordinate essential cellular processes required for survival. In pathogenicBordetellaspecies, the BvgAS TCS regulates expression of hundreds of genes, including those encoding all known protein virulence factors, and its kinase activity is essential for respiratory infection. Maintenance of BvgS kinase activity in the lower respiratory tract (LRT) depends on the function of another TCS, PlrSR. While the periplasmic venus fly-trap domains of BvgS have been implicated in responding to so-called modulating signalsin vitro(nicotinic acid and MgSO4), a role for the cytoplasmic Per-Arnt-Sim (PAS) domain in signal perception has not previously been demonstrated. By comparingB. bronchisepticastrains with mutations in the PAS domain-encoding region ofbvgSwith wild-type bacteriain vitroandin vivo, we found that although the PAS domain is not required to sense modulating signalsin vitro, it is required for the inactivation of BvgS that occurs in the absence of PlrS in the LRT of mice, suggesting that the BvgS PAS domain functions as an independent signal perception domain. Our data also indicate that the BvgS PAS domain is important for controlling absolute levels of BvgS kinase activity and the efficiency of the response to modulating signalsin vitro. Our results indicate that BvgS is capable of integrating sensory inputs from both the periplasm and the cytoplasm to control precise gene expression patterns in diverse environmental conditions.ImportanceDespite high rates of vaccination, Pertussis, a severe, highly contagious respiratory disease, caused by the bacteriumBordetella pertussis, has reemerged as a significant health threat. InBordetella pertussisand the closely related species,Bordetella bronchiseptica, activity of the BvgAS two-component regulatory system is critical for colonization of the human respiratory tract and other mammalian hosts, respectively. Here we show that the cytoplasmic PAS domain of BvgS can function as an independent signal perception domain that is capable of integrating environmental signals that influence overall BvgS activity. Our work is significant as it reveals a critical, yet previously unrecognized role, for the PAS domain in the BvgAS phosphorelay and provides a greater understanding of virulence regulation inBordetella.

scholarly journals Neither the Bvg− Phase nor thevrg6 Locus of Bordetella pertussis Is Required for Respiratory Infection in Mice

1998 ◽  
Vol 66 (6) ◽  
pp. 2762-2768 ◽  
Author(s):  
Guillermo Martinez de Tejada ◽  
Peggy A. Cotter ◽  
Ulrich Heininger ◽  
Andrew Camilli ◽  
Brian J. Akerley ◽  
...  
Keyword(s):  
Respiratory Infection ◽  
Bordetella Pertussis ◽  
Whooping Cough ◽  
Ectopic Expression ◽  
Sensory Transduction ◽  
Insertion Mutation ◽  
Wild Type ◽  
Animal Reservoir ◽  
Phenotypic Alteration

ABSTRACT In Bordetella species, the BvgAS sensory transduction system mediates an alteration between the Bvg+ phase, characterized by expression of adhesins and toxins, and the Bvg− phase, characterized by the expression of motility and coregulated phenotypes in Bordetella bronchiseptica and by the expression of vrg loci in Bordetella pertussis. Since there is no known environmental or animal reservoir for B. pertussis, the causative agent of whooping cough, it has been assumed that this phenotypic alteration must occur within the human host during infection. Consistent with this hypothesis was the observation that a B. pertussis mutant, SK6, containing a TnphoA insertion mutation in a Bvg-repressed gene (vrg6) was defective for tracheal and lung colonization in a mouse model of respiratory infection (D. T. Beattie, R. Shahin, and J. Mekalanos, Infect. Immun. 60:571–577, 1992). This result was inconsistent, however, with the observation that a Bvg+ phase-locked B. bronchiseptica mutant was indistinguishable from the wild type in its ability to establish a persistent respiratory infection in rabbits and rats (P. A. Cotter and J. F. Miller, Infect. Immun. 62:3381–3390, 1994; B. J. Akerley, P. A. Cotter, and J. F. Miller, Cell 80:611–620, 1995). To directly address the role of Bvg-mediated signal transduction in B. pertussis pathogenesis, we constructed Bvg+ and Bvg− phase-locked mutants and compared them with the wild type for their ability to colonize the respiratory tracts of mice. Our results show that the Bvg+phase of B. pertussis is necessary and sufficient for respiratory infection. By constructing a strain with a deletion in thebvgR regulatory locus, we also show that ectopic expression of Bvg− phase phenotypes decreases the efficiency of colonization, underscoring the importance of Bvg-mediated repression of gene expression in vivo. Finally, we show that the virulence defect present in strain SK6 cannot be attributed to the vrg6mutation. These data contradict an in vivo role for the Bvg− phase of B. pertussis.

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