Biology subjects

Ishmukhametov, R.

Publications and source records attributed to Ishmukhametov, R..

3 recordsLinked to original sources

bioRxiv2020

Subunit epsilon of E. coli F1Fo ATP synthase attenuates enzyme activity by modulating central stalk flexibility

F1Fo ATP synthase functions as a biological rotary generator that makes a major contribution to cellular energy production. Proton flow through the Fo motor generates rotation of the central stalk, inducing conformational changes in the F1 motor that catalyzes ATP production via flexible coupling. Here we present a range of cryo-EM structures of E. coli ATP synthase in different rotational and inhibited states observed following a 45 second incubation with 10 mM MgATP. The structures generated describe multiple changes that occur following addition of MgATP, with the inhibitory C-terminal domain of subunit {varepsilon} ({varepsilon}CTD) disassociating from the central stalk to adopt a condensed "down" conformation. The transition to the {varepsilon}CTD down state increases the torsional flexibility of the central stalk allowing its foot to rotate by [~]50{degrees}, with further flexing in the peripheral stalk enabling the c-ring to rotate by two sub-steps in the Fo motor. Truncation mutants lacking the second helix of the {varepsilon}CTD suggest that central stalk rotational flexibility is important for F1Fo ATP synthase function. Overall this study identifies the potential role played by torsional flexing within the rotor and how this could be influenced by the {varepsilon} subunit.

biophysics↗

bioRxiv2020

Swimming ghosts of the halophilic archaeon, Haloferax volcanii

Motility is seen across all domains of life1. Prokaryotes exhibit various types of motilities, such as gliding, swimming, and twitching, driven by supramolecular motility machinery composed of multiple different proteins2. In archaea only swimming motility is reported, driven by the archaellum (archaeal flagellum), a reversible rotary motor consisting of a torque-generating motor and a helical filament which acts as a propeller3,4. Unlike the bacterial flagellar motor (BFM), adenosine triphosphate (ATP) hydrolysis probably drives both motor rotation and filamentous assembly in the archaellum5,6. However, direct evidence is still lacking due to the lack of a versatile model system. Here we present a membrane-permeabilized ghost system that enables the manipulation of intracellular contents, analogous to the triton model in eukaryotic flagella7 and gliding Mycoplasma8,9. We observed high nucleotide selectivity for ATP driving motor rotation, negative cooperativity in ATP hydrolysis and the energetic requirement for at least 12 ATP molecules to be hydrolyzed per revolution of the motor. The response regulator CheY increased motor switching from counterclockwise (CCW) to clockwise (CW) rotation, which is the opposite of a previous report10. Finally, we constructed the torque-speed curve at various [ATP]s and discuss rotary models in which the archaellum has characteristics of both the BFM and F1-ATPase. Because archaea share similar cell division and chemotaxis machinery with other domains of life11,12, our ghost model will be an important tool for the exploration of the universality, diversity, and evolution of biomolecular machinery.

microbiology↗

bioRxiv2019

Visualizing movements in E. coli F1Fo ATP synthase indicates how the F1 and Fo motors are coupled

F1Fo ATP synthase functions as a biological rotary generator and makes a major contribution to cellular energy production. It is comprised of two motors that are coupled together by a central \"rotor\" and peripheral \"stator\" stalk. Proton flow through the Fo motor generates rotation of the central stalk that induces conformation changes that catalyze production of ATP in the F1 motor. Here we provide 3-4 [A] resolution cryo-EM structures of E. coli F1Fo ATP synthase in 10 mM MgADP. In addition to generating a comprehensive structural model of E. coli F1Fo ATP synthase to provide a framework to interpret mutagenesis studies, we describe a rotational sub-step of the Fo motor c-ring associated with long-range conformational changes that suggests an elegant mechanism by which the F1 and Fo motors can be coupled with minimal energy loss.

biophysics↗