Biophysics of RNA-Guided CRISPR Immunity

Loeff, Luuk; Joo, Chirlmin

doi:10.1007/978-1-4939-9726-8_9

Cited by 1 publication

(1 citation statement)

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“…Applying these criteria, the bleaching steps in each trace were determined and counted by eye by two independent researchers. This procedure was facilitated by using the fit generating function of AutoStepfinder (69). For each trace, fits displaying varying numbers of bleaching steps were created by adapting the parameters.…”

Section: Methodsmentioning

confidence: 99%

Autophosphorylation of the KaiC-like protein ArlH inhibits oligomerisation and interaction with ArlI, the motor ATPase of the archaellum

Machado

Vollmar

Schimpf

et al. 2021

Preprint

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Motile archaea are propelled by the archaellum, whose motor complex consists of the membrane protein ArlJ, the ATPase ArlI, and the ATP-binding protein ArlH. Despite its essential function and the existence of structural and biochemical data on ArlH, the role of ArlH in archaellum assembly and function remains elusive. ArlH is a structural homolog of KaiC, the central component of the cyanobacterial circadian clock. Similar to KaiC, ArlH exhibits autophosphorylation activity, which was observed for both ArlH of the euryarchaeon Pyrococcus furiosus (PfArlH) and the crenarchaeon Sulfolobus acidocaldarius (SaArlH). Using a combination of single molecule fluorescence measurements and biochemical assays, it is shown that autophosphorylation of ArlH is closely linked to the oligomeric state of ArlH bound to ArlI. These experiments also strongly suggest that ArlH is a hexamer in its functional ArlI bound state. Mutagenesis of the putative catalytic residue Glu-57 in SaArlH results in a reduced autophosphorylation activity and abolished archaellation and motility, suggesting that optimum phosphorylation activity of ArlH is essential for both archaellation and motility.

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Section: Methodsmentioning

confidence: 99%