Control of rotation of the F1FO-ATP synthase nanomotor by an inhibitory α-helix from unfolded ε or intrinsically disordered ζ and IF1 proteins

Mendoza-Hoffmann, Francisco; Zarco-Zavala, Mariel; Ortega, Raquel; García-Trejo, José J.

doi:10.1007/s10863-018-9773-9

Cited by 17 publications

(47 citation statements)

References 141 publications

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“…ATP synthases (ATPases) are a class of intrinsic membrane bound proton pumps that couple energy derived from hydrolyzing ATP to push protons uphill against a concentration gradient across the plasma membrane. There are three major classes of ATPases [113], such as archaeal A-Type ATP synthase [114,115], F-Type ATPase found in bacterial plasma membranes, mitochondrial inner membranes, and in chloroplast thylakoid membranes [116], and vacuolar-type H + -ATPase (V-ATPase) found in plasma membrane of eukaryotic specialized cells and within the membranes of organelles, such as lysosomes, endosomes, and secretory vesicles [117,118], and in some bacteria [119]. Eukaryotic vacuolar-type ATPases (V-ATPase) are the most complex (900 KDa) and the most evolved members of this protein family.…”

Section: Single-particle Cryo-em Analysis Of Membrane Proteinsmentioning

confidence: 99%

Structure Determination by Single-Particle Cryo-Electron Microscopy: Only the Sky (and Intrinsic Disorder) is the Limit

Nwanochie

Uversky

2019

IJMS

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Traditionally, X-ray crystallography and NMR spectroscopy represent major workhorses of structural biologists, with the lion share of protein structures reported in protein data bank (PDB) being generated by these powerful techniques. Despite their wide utilization in protein structure determination, these two techniques have logical limitations, with X-ray crystallography being unsuitable for the analysis of highly dynamic structures and with NMR spectroscopy being restricted to the analysis of relatively small proteins. In recent years, we have witnessed an explosive development of the techniques based on Cryo-electron microscopy (Cryo-EM) for structural characterization of biological molecules. In fact, single-particle Cryo-EM is a special niche as it is a technique of choice for the structural analysis of large, structurally heterogeneous, and dynamic complexes. Here, sub-nanometer atomic resolution can be achieved (i.e., resolution below 10 Å) via single-particle imaging of non-crystalline specimens, with accurate 3D reconstruction being generated based on the computational averaging of multiple 2D projection images of the same particle that was frozen rapidly in solution. We provide here a brief overview of single-particle Cryo-EM and show how Cryo-EM has revolutionized structural investigations of membrane proteins. We also show that the presence of intrinsically disordered or flexible regions in a target protein represents one of the major limitations of this promising technique.

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Section: Single-particle Cryo-em Analysis Of Membrane Proteinsmentioning

confidence: 99%

Structure Determination by Single-Particle Cryo-Electron Microscopy: Only the Sky (and Intrinsic Disorder) is the Limit

Nwanochie

Uversky

2019

IJMS

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“…In the active bacterial enzyme, the εCTD is proposed to adopt a condensed or ‘down’ conformation (Krah et al, 2017), and this state has been observed for isolated ε subunit from E. coli (Uhlin et al, 1997; Wilkens and Capaldi, 1998a) and Geobacillus stearothermophilus (or Bacillus PS3, hereafter termed PS3) (Yagi et al, 2007) as well as in isolated F 1 from Caldalkalibacillus thermarum (Ferguson et al, 2016). In autoinhibited states observed in crystal structures of F 1 from E. coli and PS3, the εCTDs are extended in similar ‘up’ conformations, so that a helix inserts into the F 1 central cavity and appears to block rotation of the complex by binding to both the central rotor subunit γ and several surrounding α and β subunits (Sielaff et al, 2018; Cingolani and Duncan, 2011; Mendoza-Hoffmann et al, 2018) (Figure 1—figure supplement 2). However, there is a key structural difference between these ‘up’ states: in E. coli, the εCTD consists of two helices (here termed εCTH1 and εCTH2; see Figure 1—figure supplement 2) separated by an extended loop, with each helix interacting with a different region of the γ subunit (Cingolani and Duncan, 2011; Rodgers and Wilce, 2000); whereas in PS3, εCTH1 and εCTH2 instead join to form one continuous helix (Shirakihara et al, 2015).…”

Section: Introductionmentioning

confidence: 98%

Cryo-EM reveals distinct conformations of E. coli ATP synthase on exposure to ATP

Sobti

Ishmukhametov

Bouwer

et al. 2019

eLife

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ATP synthase produces the majority of cellular energy in most cells. We have previously reported cryo-EM maps of autoinhibited E. coli ATP synthase imaged without addition of nucleotide (Sobti et al. 2016), indicating that the subunit ε engages the α, β and γ subunits to lock the enzyme and prevent functional rotation. Here we present multiple cryo-EM reconstructions of the enzyme frozen after the addition of MgATP to identify the changes that occur when this ε inhibition is removed. The maps generated show that, after exposure to MgATP, E. coli ATP synthase adopts a different conformation with a catalytic subunit changing conformation substantially and the ε C-terminal domain transitioning via an intermediate ‘half-up’ state to a condensed ‘down’ state. This work provides direct evidence for unique conformational states that occur in E. coli ATP synthase when ATP binding prevents the ε C-terminal domain from entering the inhibitory ‘up’ state.

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“…The mechanism of ion permeation control does not exist only in simple ion channels. In fact, in ATPase, which is a complicated complex, it is known that ε-protein and IF1 regulate ATPase function to regulate ion flow (35–38). Further, owing to the structural transformation of the ε subunit into the F 1 complex, which is composed of the γ and αβ subunits, the interaction of the ε subunit with the rotation of the γ subunit in the CCW direction is terminated to inhibit ion permeation via ATP degradation.…”

Section: Discussionmentioning

confidence: 99%

Function of theVibrioPomB plug region based on the stator rotation model for bacterial flagellar motor

Homma

Terashima

Koiwa

et al. 2021

Preprint

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Bacterial flagella are the only real rotational motor organs in the biological world. The spiral-shaped flagellar filaments that extend from the cell surface rotate like a screw to create a propulsive force. The base of the flagellar filament has a protein motor consisting of a stator and a rotor embedded in the membrane. The motor part has stators composed of two types of membrane subunits, PomA(MotA) and PomB(MotB), which are energy converters coupled to the ion flow that assemble around the rotor. Recently, structures of the stator, in which two molecules of MotB stuck in the center of the MotA ring made of five molecules, were reported and a model in which the MotA ring rotates with respect to MotB, which is coupled to the influx of ions, was proposed. We focused on the Vibrio PomB plug region, which has been reported to control the activation of flagellar motors. We searched for the plug region, which is the interacting region, through site-directed photo-cross-linking and disulfide cross-linking experiments. Our results demonstrated that it interacts with the extracellular short loop region of PomA, which is between transmembrane 3 and 4. Although the motor halted following cross-linking, its function was recovered with a reducing reagent that disrupted the disulfide bond. Our results support the hypothesis, which has been inferred from the stator structure, that the plug region terminates the ion inflow by stopping the rotation of the rotor.

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Control of rotation of the F1FO-ATP synthase nanomotor by an inhibitory α-helix from unfolded ε or intrinsically disordered ζ and IF1 proteins

Cited by 17 publications

References 141 publications

Structure Determination by Single-Particle Cryo-Electron Microscopy: Only the Sky (and Intrinsic Disorder) is the Limit

Structure Determination by Single-Particle Cryo-Electron Microscopy: Only the Sky (and Intrinsic Disorder) is the Limit

Cryo-EM reveals distinct conformations of E. coli ATP synthase on exposure to ATP

Function of theVibrioPomB plug region based on the stator rotation model for bacterial flagellar motor

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