A non-symmetrical p97 conformation initiates a multistep recruitment of Ufd1/Npl4

Michal Arie; Donna Matzov; Rotem Karmona; Natalia Szenkier; Ariel Stanhill; Ami Navon

doi:10.1016/j.isci.2024.110061

A non-symmetrical p97 conformation initiates a multistep recruitment of Ufd1/Npl4

Michal Arie, Donna Matzov, Rotem Karmona, Natalia Szenkier, Ariel Stanhill^*, Ami Navon^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

Abstract

In vitro experiments and cryo-EM structures of p97 and its cofactor, Ufd1/Npl4 (UN), elucidated substrate processing. Yet, the structural transitions and the related ATPase cycle upon UN binding remain unresolved. We captured two discrete conformations: One in which D1 protomers are ATP bound, while the D2 subunits are in the ADP state, presumably required for substrate engagement with the D2 pore; and a heterologous nucleotide state within the D1 ring in which only two NTDs are in the “up” ATP state that favors UN binding. Further analysis suggests that initially, UN binds p97’s non-symmetrical conformation, this association promotes a structural transition upon which five NTDs shift to an “up” state and are poised to bind ATP. The UBXL domain of Npl4 was captured bound to an NTD in the ADP state, demonstrating a conformation that may provide directionality to incoming substrate and introduce the flexibility needed for substrate processing.

Original language	English
Article number	110061
Journal	iScience
Volume	27
Issue number	6
Early online date	21 May 2024
DOIs	https://doi.org/10.1016/j.isci.2024.110061
Publication status	Published - 21 Jun 2024

Bibliographical note

We thank Dr. Nadav Elad from the electron microscopy unit for his guidance and assistance in cryo-EM data collection. Dr. Meital Kupervaser and Dr. David Morgenstern from the proteomics unit for their kind help with mass spectrometry sample handling and analysis. Dr. Harry Mark Greenblatt for setting up the workstation and his technical support. Dr. Yael Fridman Sirkis and Dr. Aharon Rabinkov from the protein analysis unit for their assistance with Biacore operation. This work was supported by Israel Science Foundation (grant #2038/17 to A.N., grant #766/20 to A.S.), Israel Cancer Association (grant #20200053 to A.S.) and the Open University of Israel Research Fund (grant #41210 to A.S.).

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© 2024

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abstract = "In vitro experiments and cryo-EM structures of p97 and its cofactor, Ufd1/Npl4 (UN), elucidated substrate processing. Yet, the structural transitions and the related ATPase cycle upon UN binding remain unresolved. We captured two discrete conformations: One in which D1 protomers are ATP bound, while the D2 subunits are in the ADP state, presumably required for substrate engagement with the D2 pore; and a heterologous nucleotide state within the D1 ring in which only two NTDs are in the “up” ATP state that favors UN binding. Further analysis suggests that initially, UN binds p97{\textquoteright}s non-symmetrical conformation, this association promotes a structural transition upon which five NTDs shift to an “up” state and are poised to bind ATP. The UBXL domain of Npl4 was captured bound to an NTD in the ADP state, demonstrating a conformation that may provide directionality to incoming substrate and introduce the flexibility needed for substrate processing.",

author = "Michal Arie and Donna Matzov and Rotem Karmona and Natalia Szenkier and Ariel Stanhill and Ami Navon",

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A non-symmetrical p97 conformation initiates a multistep recruitment of Ufd1/Npl4

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