Cryo EM structure of intact rotary H+-ATPase/synthase from Thermus thermophilus

Nakanishi, Atsuko; Kishikawa, Jun-ichi; Tamakoshi, Masatada; Mitsuoka, Kaoru; Yokoyama, Ken

doi:10.1016/j.bbabio.2018.09.242

Cited by 10 publications

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“…Single particle cryo‐electron microscopy (cryo‐EM) is a powerful technique for structure elucidation of large biomolecules and biomolecular machines, and recent technological advances have catapulted this technique to the forefront of structural biology (Kuhlbrandt, , ; Nogales & Scheres, ; Vinothkumar & Henderson, ). Cryo‐EM importantly allows for the visualization of dynamic biological processes by direct visualization of biomolecules with multiple structural states (Dong et al, ; Nakanishi, Kishikawa, Tamakoshi, Mitsuoka, & Yokoyama, ; Roh et al, ). However, to date, cryo‐EM has largely been applied to protein only or protein‐nucleic acid complexes (e.g., the ribosome and spliceosome).…”

Section: Common Methods For Rna Structure Determinationmentioning

confidence: 99%

Advances that facilitate the study of large RNA structure and dynamics by nuclear magnetic resonance spectroscopy

Zhang

Keane

2019

WIREs RNA

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The characterization of functional yet nonprotein coding (nc) RNAs has expanded the role of RNA in the cell from a passive player in the central dogma of molecular biology to an active regulator of gene expression. The misregulation of ncRNA function has been linked with a variety of diseases and disorders ranging from cancers to neurodegeneration. However, a detailed molecular understanding of how ncRNAs function has been limited; due, in part, to the difficulties associated with obtaining high-resolution structures of large RNAs. Tertiary structure determination of RNA as a whole is hampered by various technical challenges, all of which are exacerbated as the size of the RNA increases. Namely, RNAs tend to be highly flexible and dynamic molecules, which are difficult to crystallize. Biomolecular nuclear magnetic resonance (NMR) spectroscopy offers a viable alternative to determining the structure of large RNA molecules that do not readily crystallize, but is itself hindered by some technical limitations. Recently, a series of advancements have allowed the biomolecular NMR field to overcome, at least in part, some of these limitations. These advances include improvements in sample preparation strategies as well as methodological improvements. Together, these innovations pave the way for the study of ever larger RNA molecules that have important biological function.

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Section: Common Methods For Rna Structure Determinationmentioning

confidence: 99%

Advances that facilitate the study of large RNA structure and dynamics by nuclear magnetic resonance spectroscopy

Zhang

Keane

2019

WIREs RNA

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“…Recently, our group determined the three rotational state structures of Tth V/A-ATPase, based on the orientation of the central rotor subunit (Fig. 3A) [27]. The position of the central rotor subunits was different for each state, that was closely matched 120° steps which occurred during the ATP hydrolysis of V 1 .…”

Section: Single Particle Analysis Of V/a-atpasesmentioning

confidence: 99%

“…LMNG and membrane proteins form a stable detergentprotein complex at a concentration even lower than CMC due to an extremely low off rate of LMNG from the membrane protein [37]. The removal of free detergent monomers and micelles from LMNG-solubilized samples improves the image contrast dramatically and enables acquisition of highresolution data [27]. potential generated by respiration to supply ATP to the cells [15].…”

Section: Sample Preparation Of Tth V/a-atpase For Single Particle Anamentioning

confidence: 99%

“…For the V/A-ATPase, X-ray crystallography has been used to determine partial structures of the central axis [18,19], peripheral stalk [20], membrane-embedded ring of c-subunits [21], hydrophilic domain of a-subunit [22], A 3 B 3 hexamer [23], and soluble V 1 region [24]. Recent breakthroughs in structure analysis using cryogenic electron microscopy (cryo-EM) allowed us to determine the whole structure of rotary ATPases, including the F-type ATPase, V-ATPase, and V/A-ATPase [25][26][27][28][29][30][31][32][33]. In fact, the structure of the (0.34 nm) hexagonal lattice of carbon atoms with high electronic conductivity properties [41,42].…”

Section: Sample Preparation Of Tth V/a-atpase For Single Particle Anamentioning

confidence: 99%

“…Recent breakthroughs in structural studies using cryo-EM by improvements in the detector and image processing methods allowed for determination of the whole structure of rotary ATPases [13,[27][28][29][30][31][32][33]. In 2015, the different rotational states of intact rotary ATPases were first observed in both the yeast V-ATPase and F-type ATP synthase, at 6-10 Å resolution, which has enabled a more detailed understanding of the molecular mechanism of these rotary motor proteins [28,29,33].…”

Section: Single Particle Analysis Of V/a-atpasesmentioning

confidence: 99%

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Cryo-EM studies of the rotary H<sup>+</sup>-ATPase/synthase from <i>Thermus thermophilus</i>

et al. 2019

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Proton-translocating rotary ATPases couple proton influx across the membrane domain and ATP hydrolysis/ synthesis in the soluble domain through rotation of the central rotor axis against the surrounding peripheral stator apparatus. It is a significant challenge to determine the structure of rotary ATPases due to their intrinsic conformational heterogeneity and instability. Recent progress of single particle analysis of protein complexes using cryogenic electron microscopy (cryo-EM) has enabled the determination of whole rotary ATPase structures and made it possible to classify different rotational states of the enzymes at a near atomic resolution. Three cryo-EM maps corresponding to different rotational states of the V/A type H + -rotary ATPase from a bacterium Thermus thermophilus provide insights into the rotation of the whole complex, which allow us to determine the movement of each subunit during rotation. In addition, this review describes methodological developments to determine higher resolution cryo-EM structures, such as specimen preparation, to improve the image contrast of membrane proteins.ATP produced by ATP synthases is a key molecule for the metabolism of every living organism. ATP synthases belong to a family of enzymes known as rotary ATP synthases and are categorized as F-and V-type ATPases (Fig. 1A, B). They share many structural and mechanistic features, such as mechano chemical coupling of ion translocation and ATP hydrolysis/synthesis through rotation [1][2][3][4][5][6]. Reactions mediated by rotary ATPases are reversible, and consequently can function as molecular motors to synthesize or hydrolyze ATP depending on the physiological conditions [7-9].Some archaea and eubacteria contain an ATPase acting as a proton motive force-dependent ATP synthase [10,11]. Functional features and their general structures resemble F-type ATPases, whereas primary sequences of archaeal ATPases are closely related to eukaryotic V-type ATPases, which work as proton pumps depending on ATP hydrolysis in subcellular vesicles (Fig. 1C) [2,12]. The archaeal type ATPases are sometimes referred to as A-ATPases or V/A type ATPases [5,6,13], and these are regarded as an evolutionary origin of eukaryotic V-ATPases [14]. The V/A-ATPase from a thermophilic eubacterium Thermus thermophilus (Tth) is one of the best characterized rotary ATPases. The subunit composition of the Tth V/A-ATPase is similar to that of the eukaryotic enzyme; however, it has a simpler subunit structure and its physiological role is ATP synthesis in vivo, using energy from an electrochemical ATP hydrolysis/synthesis in the soluble domain of proton-translocating rotary ATPases is coupled with proton flux across the membrane domain via a rotation of the common central rotor complex against the surrounding peripheral stator apparatus. Structures of the complete V/A type H + -rotary ATPase have been revealed at a near atomic resolution level using cryogenic electron microscopy (cryo-EM). In this review article, we will describe the information about specim...

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Pharmacological Targeting of Vacuolar H+-ATPase via Subunit V1G Combats Multidrug-Resistant Cancer

Wang

Zhang

Wei

et al. 2020

Cell Chemical Biology

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Cryo EM structure of intact rotary H+-ATPase/synthase from Thermus thermophilus

Cited by 10 publications

References 0 publications

Advances that facilitate the study of large RNA structure and dynamics by nuclear magnetic resonance spectroscopy

Advances that facilitate the study of large RNA structure and dynamics by nuclear magnetic resonance spectroscopy

Cryo-EM studies of the rotary H<sup>+</sup>-ATPase/synthase from <i>Thermus thermophilus</i>

Pharmacological Targeting of Vacuolar H+-ATPase via Subunit V1G Combats Multidrug-Resistant Cancer

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