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Structural and mechanistic basis for protein glutamylation by the kinase fold

Adam Osinski, Miles H. Black, Krzysztof Pawłowski, Zhe Chen, Yang Li, Vincent S. Tagliabracci
doi: https://doi.org/10.1101/2021.04.13.439722
Adam Osinski
1Department of Molecular Biology, University of Texas Southwestern Medical Center; Dallas, TX 75390, USA
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Miles H. Black
1Department of Molecular Biology, University of Texas Southwestern Medical Center; Dallas, TX 75390, USA
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Krzysztof Pawłowski
1Department of Molecular Biology, University of Texas Southwestern Medical Center; Dallas, TX 75390, USA
2Institute of Biology, Warsaw University of Life Sciences; Warsaw, 02-787, Poland
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Zhe Chen
3Department of Biophysics, University of Texas Southwestern Medical Center; Dallas, TX 75390, USA
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Yang Li
3Department of Biophysics, University of Texas Southwestern Medical Center; Dallas, TX 75390, USA
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  • For correspondence: Yang2.Li@UTSouthwestern.edu Vincent.Tagliabracci@UTSouthwestern.edu
Vincent S. Tagliabracci
1Department of Molecular Biology, University of Texas Southwestern Medical Center; Dallas, TX 75390, USA
4Harold C. Simmons Comprehensive Cancer Center, University of Texas Southwestern Medical Center; Dallas, TX, 75390, USA
5Hamon Center for Regenerative Science and Medicine, University of Texas Southwestern Medical Center; Dallas, TX, 75390, USA
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  • For correspondence: Yang2.Li@UTSouthwestern.edu Vincent.Tagliabracci@UTSouthwestern.edu
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Summary

The kinase domain transfers phosphate from ATP to substrates. However, the Legionella effector SidJ adopts a kinase fold yet catalyzes calmodulin (CaM)-dependent glutamylation to inactivate the SidE ubiquitin ligases. The structural and mechanistic basis in which the kinase domain catalyzes protein glutamylation is unknown. Here we present cryo-EM reconstructions of SidJ:CaM:SidE reaction intermediate complexes. We show that the kinase-like active site of SidJ adenylates an active site Glu in SidE resulting in the formation of a stable reaction intermediate complex. An insertion in the catalytic loop of the kinase domain positions the donor Glu near the acyl-adenylate for peptide bond formation. Our structural analysis led us to discover that the SidJ paralog SdjA is a glutamylase that differentially regulates the SidE-ligases during Legionella infection. Our results uncover the structural and mechanistic basis in which the kinase fold catalyzes non-ribosomal amino acid ligations and reveal an unappreciated level of SidE-family regulation.

Competing Interest Statement

The authors have declared no competing interest.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-ND 4.0 International license.
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Posted April 13, 2021.
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Structural and mechanistic basis for protein glutamylation by the kinase fold
Adam Osinski, Miles H. Black, Krzysztof Pawłowski, Zhe Chen, Yang Li, Vincent S. Tagliabracci
bioRxiv 2021.04.13.439722; doi: https://doi.org/10.1101/2021.04.13.439722
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Structural and mechanistic basis for protein glutamylation by the kinase fold
Adam Osinski, Miles H. Black, Krzysztof Pawłowski, Zhe Chen, Yang Li, Vincent S. Tagliabracci
bioRxiv 2021.04.13.439722; doi: https://doi.org/10.1101/2021.04.13.439722

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