scholarly journals Structural photoactivation of a full-length bacterial phytochrome

2016 ◽  
Vol 2 (8) ◽  
pp. e1600920 ◽  
Author(s):  
Alexander Björling ◽  
Oskar Berntsson ◽  
Heli Lehtivuori ◽  
Heikki Takala ◽  
Ashley J. Hughes ◽  
...  
Keyword(s):  
Time Scales ◽  
Conformational Changes ◽  
Structural Changes ◽  
Deinococcus Radiodurans ◽  
Full Length ◽  
Photon Absorption ◽  
Signaling Mechanism ◽  
Time Resolved ◽  
Binding Domains ◽  
Sensor Proteins

Phytochromes are light sensor proteins found in plants, bacteria, and fungi. They function by converting a photon absorption event into a conformational signal that propagates from the chromophore through the entire protein. However, the structure of the photoactivated state and the conformational changes that lead to it are not known. We report time-resolved x-ray scattering of the full-length phytochrome from Deinococcus radiodurans on micro- and millisecond time scales. We identify a twist of the histidine kinase output domains with respect to the chromophore-binding domains as the dominant change between the photoactivated and resting states. The time-resolved data further show that the structural changes up to the microsecond time scales are small and localized in the chromophore-binding domains. The global structural change occurs within a few milliseconds, coinciding with the formation of the spectroscopic meta-Rc state. Our findings establish key elements of the signaling mechanism of full-length bacterial phytochromes.

scholarly journals On the (un)coupling of the chromophore, tongue interactions, and overall conformation in a bacterial phytochrome

2018 ◽  
Vol 293 (21) ◽  
pp. 8161-8172 ◽  
Author(s):  
Heikki Takala ◽  
Heli K. Lehtivuori ◽  
Oskar Berntsson ◽  
Ashley Hughes ◽  
Rahul Nanekar ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Structural Changes ◽  
Deinococcus Radiodurans ◽  
Red Light ◽  
Light Responses ◽  
Structural Mechanism ◽  
Weakly Coupled ◽  
Activity State ◽  
Conserved Tyrosine

Phytochromes are photoreceptors in plants, fungi, and various microorganisms and cycle between metastable red light–absorbing (Pr) and far-red light–absorbing (Pfr) states. Their light responses are thought to follow a conserved structural mechanism that is triggered by isomerization of the chromophore. Downstream structural changes involve refolding of the so-called tongue extension of the phytochrome-specific GAF-related (PHY) domain of the photoreceptor. The tongue is connected to the chromophore by conserved DIP and PRXSF motifs and a conserved tyrosine, but the role of these residues in signal transduction is not clear. Here, we examine the tongue interactions and their interplay with the chromophore by substituting the conserved tyrosine (Tyr263) in the phytochrome from the extremophile bacterium Deinococcus radiodurans with phenylalanine. Using optical and FTIR spectroscopy, X-ray solution scattering, and crystallography of chromophore-binding domain (CBD) and CBD–PHY fragments, we show that the absence of the Tyr263 hydroxyl destabilizes the β-sheet conformation of the tongue. This allowed the phytochrome to adopt an α-helical tongue conformation regardless of the chromophore state, hence distorting the activity state of the protein. Our crystal structures further revealed that water interactions are missing in the Y263F mutant, correlating with a decrease of the photoconversion yield and underpinning the functional role of Tyr263 in phytochrome conformational changes. We propose a model in which isomerization of the chromophore, refolding of the tongue, and globular conformational changes are represented as weakly coupled equilibria. The results also suggest that the phytochromes have several redundant signaling routes.

scholarly journals Light-induced structural changes in a full-length cyanobacterial phytochrome probed by time-resolved X-ray scattering

2019 ◽  
Vol 2 (1) ◽  
Author(s):  
Derren J. Heyes ◽  
Samantha J. O. Hardman ◽  
Martin N. Pedersen ◽  
Joyce Woodhouse ◽  
Eugenio De La Mora ◽  
...  
Keyword(s):  
Structural Changes ◽  
Full Length ◽  
Time Resolved ◽  
X Ray ◽  
X Ray Scattering ◽  
Ray Scattering

Stroboscopic neutron diffraction from spatially modulated systems

2003 ◽  
Vol 218 (2) ◽  
Author(s):  
G. Eckold ◽  
H. Gibhardt ◽  
D. Caspary ◽  
P. Elter ◽  
K. Elisbihani
Keyword(s):  
Neutron Diffraction ◽  
Time Scales ◽  
Structural Changes ◽  
Structural Response ◽  
Relaxation Processes ◽  
Time Resolved ◽  
Transient States ◽  
Underlying Mechanisms ◽  
Lock In ◽  
Kinetics Of

AbstractThe combination of stroboscopic techniques and neutron diffraction can be used to study the kinetics of structural changes in condensed matter on a microscopic level. Transient states may be identified and characterized on time-scales down to the microsecond regime. Hence, valuable information about the underlying mechanisms can be obtained from time-resolved experiments. Particularly interesting subjects for this type of investigation are spatially modulated systems which undergo phase transitions or phase separation. The potential of stroboscopic neutron diffraction is demonstrated using three different examples in which samples are periodically perturbed by the variation of temperature, mechanical stress or electric field and their structural response is characterized by time-resolved diffraction. Spinodal decomposition in ionic crystals of the silver-alkali halide type is shown to be dominated by two different processes on different time-scales. The stress-induced phase transition into the incommensurate phase of quartz involves relaxation processes which are reflected by different kinetic behaviours of Bragg peaks and satellite reflections, respectively. Finally, metastable transient states are observed during the field induced lock-in transition in ferroelectric Rb

scholarly journals Unidirectional ion transport mechanism of a light-driven chloride pump revealed using X-ray free electron lasers

2021 ◽  
Author(s):  
Toshiaki Hosaka ◽  
Takashi Nomura ◽  
Minoru Kubo ◽  
Takanori Nakane ◽  
Luo Fangjia ◽  
...  
Keyword(s):  
Ion Transport ◽  
Conformational Changes ◽  
Structural Changes ◽  
Ion Transfer ◽  
Ion Release ◽  
Time Resolved ◽  
Iodide Ions ◽  
Lateral Displacements ◽  
Extracellular Side ◽  
Cytoplasmic Side

Light-driven chloride-pumping rhodopsins actively transport anions, including various halide ions, across cell membranes. Recent studies using time-resolved serial femtosecond crystallography (TR-SFX) have uncovered the structural changes and ion transfer mechanisms in light-driven cation-pumping rhodopsins. However, the mechanism by which the conformational changes pump an anion to achieve unidirectional ion transport, from the extracellular side to the cytoplasmic side, in anion-pumping rhodopsins remains enigmatic. We have collected TR-SFX data of Nonlabens marinus rhodopsin-3 (NM-R3), derived from a marine flavobacterium, at 10 µs and 1 ms time-points after photoexcitation. Our structural analysis reveals the conformational alterations during ion transfer and after ion release. Movements of the retinal chromophore initially displace a conserved tryptophan to the cytoplasmic side of NM-R3, accompanied with a slight shift of the halide ion bound to the retinal. After ion release, the inward movements of helix C and helix G and the lateral displacements of the retinal block access to the extracellular side of NM-R3. Anomalous signal data have also been obtained from NM-R3 crystals containing iodide ions. The anomalous density maps provide insight into the halide binding site for ion transfer in NM-R3.

scholarly journals Insulin-Induced Conformational Changes in the Full-Length Insulin Receptor: Structural Insights Gained from Molecular Modeling Analyses

2020 ◽  
Author(s):  
Yong Xiao Yang ◽  
Peng Li ◽  
Pan Wang ◽  
Bao Ting Zhu
Keyword(s):  
Diabetes Mellitus ◽  
Molecular Modeling ◽  
Insulin Receptor ◽  
Conformational Changes ◽  
Structural Changes ◽  
Conformational Transition ◽  
Structural Alignment ◽  
Full Length ◽  
New Drugs ◽  
Complex Dynamic

ABSTRACTInsulin receptor plays an important role in regulation of energy metabolism. Dysfunction of insulin receptor (IR) can lead to many disease states, such as diabetes mellitus. Deciphering the complex dynamic structure of human IR and its mechanism of activation would greatly aid in understanding IR-mediated signaling pathways and in particular, in designing new drugs (including nonpeptidal insulin analogs) to treat diabetes mellitus. Experimental evidence about IR structure has been gradually obtained by biologists over the past three decades. Based on the available experimental structures of IR in different states, here we employ molecular modeling approach to construct the full-length IR structures in different states and model its structural and conformational changes during insulin-induced IR activation. Several key possible intermediate states are constructed based on structural alignment, rotation and computational modeling. Based on the structures of the full-length IR in different states, it appears that there are two possible conformational transition pathways: one is symmetric, and the other one is asymmetric. Structural changes and motions of different domains of the full-length IR along the pathways are analyzed. The role of insulin binding to IR in facilitating the conformational transition of the receptor is modeled. Information and insights derived from our present structural modeling analyses may aid in understanding the complex dynamic, structural and conformational changes during the process of IR activation.

scholarly journals Photocage-initiated time-resolved solution X-ray scattering investigation of protein dimerization

IUCrJ ◽  
2018 ◽  
Vol 5 (6) ◽  
pp. 667-672 ◽  
Author(s):  
Inokentijs Josts ◽  
Stephan Niebling ◽  
Yunyun Gao ◽  
Matteo Levantino ◽  
Henning Tidow ◽  
...  
Keyword(s):  
Small Molecule ◽  
Conformational Changes ◽  
Structural Changes ◽  
Time Resolved ◽  
Protein Dimerization ◽  
X Ray ◽  
Single Turnover ◽  
X Ray Scattering ◽  
Photoactivatable Proteins ◽  
Ray Scattering

This work demonstrates a new method for investigating time-resolved structural changes in protein conformation and oligomerization via photocage-initiated time-resolved X-ray solution scattering by observing the ATP-driven dimerization of the MsbA nucleotide-binding domain. Photocaged small molecules allow the observation of single-turnover reactions of non-naturally photoactivatable proteins. The kinetics of the reaction can be derived from changes in X-ray scattering associated with ATP-binding and subsequent dimerization. This method can be expanded to any small-molecule-driven protein reaction with conformational changes traceable by X-ray scattering where the small molecule can be photocaged.

scholarly journals Time-resolved serial femtosecond crystallography reveals early structural changes in channelrhodopsin

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Kazumasa Oda ◽  
Takashi Nomura ◽  
Takanori Nakane ◽  
Keitaro Yamashita ◽  
Keiichi Inoue ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Structural Changes ◽  
Local Deformation ◽  
Proton Donor ◽  
Time Resolved ◽  
Molecular Events ◽  
Channel Opening ◽  
Ion Conducting ◽  
And Shifts ◽  
Opening And Closing

Channelrhodopsins (ChRs) are microbial light-gated ion channels utilized in optogenetics to control neural activity with light . Light absorption causes retinal chromophore isomerization and subsequent protein conformational changes visualized as optically distinguished intermediates, coupled with channel opening and closing. However, the detailed molecular events underlying channel gating remain unknown. We performed time-resolved serial femtosecond crystallographic analyses of ChR by using an X-ray free electron laser, which revealed conformational changes following photoactivation. The isomerized retinal adopts a twisted conformation and shifts toward the putative internal proton donor residues, consequently inducing an outward shift of TM3, as well as a local deformation in TM7. These early conformational changes in the pore-forming helices should be the triggers that lead to opening of the ion conducting pore.

scholarly journals Structural changes of tailless bacteriophage ΦX174 during penetration of bacterial cell walls

2017 ◽  
Vol 114 (52) ◽  
pp. 13708-13713 ◽  
Author(s):  
Yingyuan Sun ◽  
Aaron P. Roznowski ◽  
Joshua M. Tokuda ◽  
Thomas Klose ◽  
Alexander Mauney ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Cell Walls ◽  
Structural Changes ◽  
The Other ◽  
Host Bacterium ◽  
Time Resolved ◽  
Viral Attachment ◽  
X Ray ◽  
X Ray Scattering ◽  
Cryo Electron Microscopy

Unlike tailed bacteriophages, which use a preformed tail for transporting their genomes into a host bacterium, the ssDNA bacteriophage ΦX174 is tailless. Using cryo-electron microscopy and time-resolved small-angle X-ray scattering, we show that lipopolysaccharides (LPS) form bilayers that interact with ΦX174 at an icosahedral fivefold vertex and induce single-stranded (ss) DNA genome ejection. The structures of ΦX174 complexed with LPS have been determined for the pre- and post-ssDNA ejection states. The ejection is initiated by the loss of the G protein spike that encounters the LPS, followed by conformational changes of two polypeptide loops on the major capsid F proteins. One of these loops mediates viral attachment, and the other participates in making the fivefold channel at the vertex contacting the LPS.

scholarly journals Photoactivation of Drosophila melanogaster cryptochrome through sequential conformational transitions

2019 ◽  
Vol 5 (7) ◽  
pp. eaaw1531 ◽  
Author(s):  
Oskar Berntsson ◽  
Ryan Rodriguez ◽  
Léocadie Henry ◽  
Matthijs R. Panman ◽  
Ashley J. Hughes ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Conformational Changes ◽  
Structural Changes ◽  
Structural Response ◽  
Biochemical Activity ◽  
Time Resolved ◽  
X Ray ◽  
Molecular Events ◽  
Carboxyl Terminal ◽  
Dynamics Simulations

Cryptochromes are blue-light photoreceptor proteins, which provide input to circadian clocks. The cryptochrome from Drosophila melanogaster (DmCry) modulates the degradation of Timeless and itself. It is unclear how light absorption by the chromophore and the subsequent redox reactions trigger these events. Here, we use nano- to millisecond time-resolved x-ray solution scattering to reveal the light-activated conformational changes in DmCry and the related (6-4) photolyase. DmCry undergoes a series of structural changes, culminating in the release of the carboxyl-terminal tail (CTT). The photolyase has a simpler structural response. We find that the CTT release in DmCry depends on pH. Mutation of a conserved histidine, important for the biochemical activity of DmCry, does not affect transduction of the structural signal to the CTT. Instead, molecular dynamics simulations suggest that it stabilizes the CTT in the resting-state conformation. Our structural photocycle unravels the first molecular events of signal transduction in an animal cryptochrome.

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