scholarly journals Topography and motion of the acid-sensing ion channel intracellular domains

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Tyler Couch ◽  
Kyle Berger ◽  
Dana L Kneisley ◽  
Tyler W McCullock ◽  
Paul Kammermeier ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Fluorescent Proteins ◽  
Topographic Map ◽  
Relative Movement ◽  
Extracellular Acidification ◽  
X Ray ◽  
Transmembrane Segments ◽  
Lifetime Imaging ◽  
Acid Sensing Ion Channels ◽  
Intracellular Domains

Acid-sensing ion channels (ASICs) are trimeric cation-selective channels activated by decreases in extracellular pH. The intracellular N and C terminal tails of ASIC1 influence channel gating, trafficking, and signaling in ischemic cell death. Despite several x-ray and cryo-EM structures of the extracellular and transmembrane segments of ASIC1, these important intracellular tails remain unresolved. Here we describe the coarse topography of the chicken ASIC1 intracellular domains determined by FRET, measured using either fluorescent lifetime imaging or patch clamp fluorometry. We find the C terminal tail projects into the cytosol by approximately 35 Å and that the N and C tail from the same subunits are closer than adjacent subunits. Using pH-insensitive fluorescent proteins, we fail to detect any relative movement between the N and C tails upon extracellular acidification but do observe axial motions of the membrane proximal segments towards the plasma membrane. Taken together, our study furnishes a coarse topographic map of the ASIC intracellular domains while providing directionality and context to intracellular conformational changes induced by extracellular acidification.

scholarly journals Topography and motion of the acid-sensing ion channel intracellular domains

2021 ◽  
Author(s):  
Tyler Couch ◽  
Kyle Berger ◽  
Dana L. Kneisley ◽  
Tyler W. McCullock ◽  
Paul J. Kammermeier ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Fluorescent Proteins ◽  
Topographic Map ◽  
Relative Movement ◽  
Extracellular Acidification ◽  
X Ray ◽  
Transmembrane Segments ◽  
Lifetime Imaging ◽  
Acid Sensing Ion Channels ◽  
Intracellular Domains

AbstractAcid-sensing ion channels (ASICs) are trimeric cation-selective channels activated by decreases in extracellular pH. The intracellular N and C terminal tails of ASIC1 influence channel gating, trafficking, and signaling in ischemic cell death. Despite several x-ray and cryo-EM structures of the extracellular and transmembrane segments of ASIC1, these important intracellular tails remain unresolved. Here we describe the coarse topography of the cASIC1 intracellular domains determined by FRET, measured using either fluorescent lifetime imaging or patch clamp fluorometry. We find the C terminal tail projects into the cytosol by approximately 35 Å and that the N and proximal segment of the C tail from the same subunits are closer than adjacent subunits. Using pH-insensitive fluorescent proteins, we fail to detect any relative movement between the N and C tails upon extracellular acidification but do observe axial motions of the membrane proximal segments towards the plasma membrane. Taken together, our study furnishes a coarse topographic map of the ASIC intracellular domains while providing directionality and context to intracellular conformational changes induced by extracellular acidification.

scholarly journals Unusual Conformational Changes in 6-Hydroxymethyl-7,8-dihydropterin Pyrophosphokinase as Revealed by X-ray Crystallography and NMR

2001 ◽  
Vol 276 (43) ◽  
pp. 40274-40281 ◽  
Author(s):  
Bing Xiao ◽  
Genbin Shi ◽  
Jinhai Gao ◽  
Jaroslaw Blaszczyk ◽  
Qin Liu ◽  
...  
Keyword(s):  
Conformational Changes ◽  
X Ray ◽  
X Ray Crystallography

X-ray photoemission studies of the interaction of metals and metal ions with DNA

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Esha Mishra ◽  
Subrata Majumder ◽  
Shikha Varma ◽  
Peter A. Dowben
Keyword(s):  
Heavy Metal ◽  
Metal Ions ◽  
Conformational Changes ◽  
Photoelectron Spectroscopy ◽  
X Ray ◽  
Surface Sensitivity ◽  
Metal Interaction ◽  
The Status ◽  
Sensitivity And Selectivity ◽  
Insight Into

Abstract X-ray Photoelectron Spectroscopy (XPS) has been used to study the interactions of heavy metal ions with DNA with some success. Surface sensitivity and selectivity of XPS are advantageous for identifying and characterizing the chemical and elemental structure of the DNA to metal interaction. This review summarizes the status of what amounts to a large part of the photoemission investigations of biomolecule interactions with metals and offers insight into the mechanism for heavy metal-bio interface interactions. Specifically, it is seen that metal interaction with DNA results in conformational changes in the DNA structure.

The kinetics of conformational changes of α2 -macroglobulin determined by time resolved X-ray solution scattering

FEBS Letters ◽  
1994 ◽  
Vol 337 (2) ◽  
pp. 171-174 ◽  
Author(s):  
Hideo Arakawa ◽  
Takuji Urisaka ◽  
Hirotsugu Tsuruta ◽  
Yoshiyuki Amemiya ◽  
Hiroshi Kihara ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Time Resolved ◽  
X Ray ◽  
Α2 Macroglobulin ◽  
Kinetics Of

scholarly journals Predicting data quality in biological X-ray solution scattering

2018 ◽  
Vol 74 (8) ◽  
pp. 727-738
Author(s):  
Chenzheng Wang ◽  
Yuexia Lin ◽  
Devin Bougie ◽  
Richard E. Gillilan
Keyword(s):  
Conformational Changes ◽  
Sample Path ◽  
Flow Time ◽  
Photon Flux ◽  
Detector Efficiency ◽  
Time Resolved ◽  
Short Sample ◽  
X Ray ◽  
Path Lengths ◽  
Flux Energy

Biological small-angle X-ray solution scattering (BioSAXS) is now widely used to gain information on biomolecules in the solution state. Often, however, it is not obvious in advance whether a particular sample will scatter strongly enough to give useful data to draw conclusions under practically achievable solution conditions. Conformational changes that appear to be large may not always produce scattering curves that are distinguishable from each other at realistic concentrations and exposure times. Emerging technologies such as time-resolved SAXS (TR-SAXS) pose additional challenges owing to small beams and short sample path lengths. Beamline optics vary in brilliance and degree of background scatter, and major upgrades and improvements to sources promise to expand the reach of these methods. Computations are developed to estimate BioSAXS sample intensity at a more detailed level than previous approaches, taking into account flux, energy, sample thickness, window material, instrumental background, detector efficiency, solution conditions and other parameters. The results are validated with calibrated experiments using standard proteins on four different beamlines with various fluxes, energies and configurations. The ability of BioSAXS to statistically distinguish a variety of conformational movements under continuous-flow time-resolved conditions is then computed on a set of matched structure pairs drawn from the Database of Macromolecular Motions (http://molmovdb.org). The feasibility of experiments is ranked according to sample consumption, a quantity that varies by over two orders of magnitude for the set of structures. In addition to photon flux, the calculations suggest that window scattering and choice of wavelength are also important factors given the short sample path lengths common in such setups.

scholarly journals Microchannel Fabrication Using A Photo-Patternable Adhesive Material for Recording Conformational Changes of KcsA Channel with the Diffracted X-ray Tracking Method

Proceedings ◽  
2018 ◽  
Vol 2 (13) ◽  
pp. 972 ◽  
Author(s):  
Yoshikazu Hirai ◽  
Yasuaki Mori ◽  
Tomoki Tabuchi ◽  
Hirofumi Shimizu ◽  
Toshiyuki Tsuchiya ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Diffraction Spot ◽  
High Resolution Imaging ◽  
Potassium Ion Channel ◽  
Adhesive Material ◽  
Tracking Method ◽  
X Ray ◽  
Sin Membrane ◽  
Resolution Imaging ◽  
Gold Nanocrystal

Diffracted X-ray tracking (DXT) method can trace conformational changes of KcsA potassium ion channel during gating by recording position of diffraction spot from a gold nanocrystal attached to the channel as a movie. For high-resolution imaging under controlled microenvironments for KcsA channels, we report a microfluidic device consisting of two SiN membrane windows bonded with a photo patternable adhesive material. The reduced signal-to-background ratio as well as suitable adhesive material thickness for the microchannel are discussed in the experiment at the synchrotron radiation facility.

scholarly journals Proteasomal conformation controls unfolding ability

2021 ◽  
Vol 118 (25) ◽  
pp. e2101004118
Author(s):  
Julianna R. Cresti ◽  
Abramo J. Manfredonia ◽  
Christopher E. Bragança ◽  
Joseph A. Boscia ◽  
Christina M. Hurley ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Fluorescent Proteins ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Conformational State ◽  
26S Proteasome ◽  
Eukaryotic Cells ◽  
Ubiquitinated Protein ◽  
Equilibrium Conditions ◽  
Substrate Processing

The 26S proteasome is the macromolecular machine responsible for the bulk of protein degradation in eukaryotic cells. As it degrades a ubiquitinated protein, the proteasome transitions from a substrate-accepting conformation (s1) to a set of substrate-processing conformations (s3 like), each stabilized by different intramolecular contacts. Tools to study these conformational changes remain limited, and although several interactions have been proposed to be important for stabilizing the proteasome’s various conformations, it has been difficult to test these directly under equilibrium conditions. Here, we describe a conformationally sensitive Förster resonance energy transfer assay, in which fluorescent proteins are fused to Sem1 and Rpn6, which are nearer each other in substrate-processing conformations than in the substrate-accepting conformation. Using this assay, we find that two sets of interactions, one involving Rpn5 and another involving Rpn2, are both important for stabilizing substrate-processing conformations. Mutations that disrupt these interactions both destabilize substrate-processing conformations relative to the substrate-accepting conformation and diminish the proteasome’s ability to successfully unfold and degrade hard-to-unfold substrates, providing a link between the proteasome’s conformational state and its unfolding ability.

scholarly journals Kinetic analysis of ASIC1a delineates conformational signaling from proton-sensing domains to the channel gate

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Sabrina Vullo ◽  
Nicolas Ambrosio ◽  
Jan P Kucera ◽  
Olivier Bignucolo ◽  
Stephan Kellenberger
Keyword(s):  
Ion Channels ◽  
Kinetic Analysis ◽  
Conformational Changes ◽  
Transmembrane Helix ◽  
Activation Mechanism ◽  
Pain Sensation ◽  
Channel Activation ◽  
Channel Pore ◽  
Acid Sensing Ion Channels ◽  
Channel Gate

Acid-sensing ion channels (ASICs) are neuronal Na+ channels that are activated by a drop in pH. Their established physiological and pathological roles, involving fear behaviors, learning, pain sensation and neurodegeneration after stroke, make them promising targets for future drugs. Currently, the ASIC activation mechanism is not understood. Here we used voltage-clamp fluorometry (VCF) combined with fluorophore-quencher pairing to determine the kinetics and direction of movements. We show that conformational changes with the speed of channel activation occur close to the gate and in more distant extracellular sites, where they may be driven by local protonation events. Further, we provide evidence for fast conformational changes in a pathway linking protonation sites to the channel pore, in which an extracellular interdomain loop interacts via aromatic residue interactions with the upper end of a transmembrane helix and would thereby open the gate.

scholarly journals Directionality of light absorption and emission in representative fluorescent proteins

2020 ◽  
Vol 117 (51) ◽  
pp. 32395-32401
Author(s):  
Jitka Myšková ◽  
Olga Rybakova ◽  
Jiří Brynda ◽  
Petro Khoroshyy ◽  
Alexey Bondar ◽  
...  
Keyword(s):  
Light Wave ◽  
Fluorescent Proteins ◽  
Dipole Moments ◽  
Biological Processes ◽  
Transition Dipole ◽  
X Ray ◽  
Emission Transition ◽  
Transition Dipole Moments ◽  
Fluorescent Molecules ◽  
Incoming Light

Fluorescent molecules are like antennas: The rate at which they absorb light depends on their orientation with respect to the incoming light wave, and the apparent intensity of their emission depends on their orientation with respect to the observer. However, the directions along which the most important fluorescent molecules in biology, fluorescent proteins (FPs), absorb and emit light are generally not known. Our optical and X-ray investigations of FP crystals have now allowed us to determine the molecular orientations of the excitation and emission transition dipole moments in the FPs mTurquoise2, eGFP, and mCherry, and the photoconvertible FP mEos4b. Our results will allow using FP directionality in studies of molecular and biological processes, but also in development of novel bioengineering and bioelectronics applications.

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