Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser

Rebecca J. Jernigan; Dhenugen Logeswaran; Diandra Doppler; Nirupa Nagaratnam; Mukul Sonker; Jay How Yang; Gihan Ketawala; Jose M. Martin-Garcia; Megan L. Shelby; Thomas D. Grant; Valerio Mariani; Alexandra Tolstikova; Michelle Z. Sheikh; Mimi Cho Yung; Matthew A. Coleman; Sahba Zaare; Emily K. Kaschner; Mohammad Towshif Rabbani; Reza Nazari; Michele A. Zacks; Brandon Hayes; Raymond G. Sierra; Mark S. Hunter; Stella Lisova; Alexander Batyuk; Christopher Kupitz; Sebastien Boutet; Debra T. Hansen; Richard A. Kirian; Marius Schmidt; Raimund Fromme; Matthias Frank; Alexandra Ros; Julian J.L. Chen; Sabine Botha; Petra Fromme

doi:10.1016/j.str.2022.12.009

Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser

Rebecca J. Jernigan, Dhenugen Logeswaran, Diandra Doppler, Nirupa Nagaratnam, Mukul Sonker, Jay How Yang, Gihan Ketawala, Jose M. Martin-Garcia, Megan L. Shelby, Thomas D. Grant, Valerio Mariani, Alexandra Tolstikova, Michelle Z. Sheikh, Mimi Cho Yung, Matthew A. Coleman, Sahba Zaare, Emily K. Kaschner, Mohammad Towshif Rabbani, Reza Nazari, Michele A. ZacksBrandon Hayes, Raymond G. Sierra, Mark S. Hunter, Stella Lisova, Alexander Batyuk, Christopher Kupitz, Sebastien Boutet, Debra T. Hansen, Richard A. Kirian, Marius Schmidt, Raimund Fromme, Matthias Frank, Alexandra Ros, Julian J.L. Chen, Sabine Botha, Petra Fromme

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

5 Scopus citations

Abstract

NendoU from SARS-CoV-2 is responsible for the virus's ability to evade the innate immune system by cleaving the polyuridine leader sequence of antisense viral RNA. Here we report the room-temperature structure of NendoU, solved by serial femtosecond crystallography at an X-ray free-electron laser to 2.6 Å resolution. The room-temperature structure provides insight into the flexibility, dynamics, and other intrinsic properties of NendoU, with indications that the enzyme functions as an allosteric switch. Functional studies examining cleavage specificity in solution and in crystals support the uridine-purine cleavage preference, and we demonstrate that enzyme activity is fully maintained in crystal form. Optimizing the purification of NendoU and identifying suitable crystallization conditions set the benchmark for future time-resolved serial femtosecond crystallography studies. This could advance the design of antivirals with higher efficacy in treating coronaviral infections, since drugs that block allosteric conformational changes are less prone to drug resistance.

Original language	English (US)
Pages (from-to)	138-151.e5
Journal	Structure
Volume	31
Issue number	2
DOIs	https://doi.org/10.1016/j.str.2022.12.009
State	Published - Feb 2 2023

Keywords

COVID-19
NSP15
NendoU
SARS-CoV-2
X-ray free-electron laser
serial femtosecond crystallography

ASJC Scopus subject areas

Structural Biology
Molecular Biology

Access to Document

10.1016/j.str.2022.12.009

Cite this

Jernigan, R. J., Logeswaran, D., Doppler, D., Nagaratnam, N., Sonker, M., Yang, J. H., Ketawala, G., Martin-Garcia, J. M., Shelby, M. L., Grant, T. D., Mariani, V., Tolstikova, A., Sheikh, M. Z., Yung, M. C., Coleman, M. A., Zaare, S., Kaschner, E. K., Rabbani, M. T., Nazari, R., ... Fromme, P. (2023). Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser. Structure, 31(2), 138-151.e5. https://doi.org/10.1016/j.str.2022.12.009

Jernigan, RJ, Logeswaran, D, Doppler, D, Nagaratnam, N, Sonker, M, Yang, JH, Ketawala, G, Martin-Garcia, JM, Shelby, ML, Grant, TD, Mariani, V, Tolstikova, A, Sheikh, MZ, Yung, MC, Coleman, MA, Zaare, S, Kaschner, EK, Rabbani, MT, Nazari, R, Zacks, MA, Hayes, B, Sierra, RG, Hunter, MS, Lisova, S, Batyuk, A, Kupitz, C, Boutet, S, Hansen, DT , Kirian, RA, Schmidt, M, Fromme, R, Frank, M, Ros, A , Chen, JJL, Botha, S & Fromme, P 2023, 'Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser', Structure, vol. 31, no. 2, pp. 138-151.e5. https://doi.org/10.1016/j.str.2022.12.009

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title = "Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser",

abstract = "NendoU from SARS-CoV-2 is responsible for the virus's ability to evade the innate immune system by cleaving the polyuridine leader sequence of antisense viral RNA. Here we report the room-temperature structure of NendoU, solved by serial femtosecond crystallography at an X-ray free-electron laser to 2.6 {\AA} resolution. The room-temperature structure provides insight into the flexibility, dynamics, and other intrinsic properties of NendoU, with indications that the enzyme functions as an allosteric switch. Functional studies examining cleavage specificity in solution and in crystals support the uridine-purine cleavage preference, and we demonstrate that enzyme activity is fully maintained in crystal form. Optimizing the purification of NendoU and identifying suitable crystallization conditions set the benchmark for future time-resolved serial femtosecond crystallography studies. This could advance the design of antivirals with higher efficacy in treating coronaviral infections, since drugs that block allosteric conformational changes are less prone to drug resistance.",

keywords = "COVID-19, NSP15, NendoU, SARS-CoV-2, X-ray free-electron laser, serial femtosecond crystallography",

author = "Jernigan, {Rebecca J.} and Dhenugen Logeswaran and Diandra Doppler and Nirupa Nagaratnam and Mukul Sonker and Yang, {Jay How} and Gihan Ketawala and Martin-Garcia, {Jose M.} and Shelby, {Megan L.} and Grant, {Thomas D.} and Valerio Mariani and Alexandra Tolstikova and Sheikh, {Michelle Z.} and Yung, {Mimi Cho} and Coleman, {Matthew A.} and Sahba Zaare and Kaschner, {Emily K.} and Rabbani, {Mohammad Towshif} and Reza Nazari and Zacks, {Michele A.} and Brandon Hayes and Sierra, {Raymond G.} and Hunter, {Mark S.} and Stella Lisova and Alexander Batyuk and Christopher Kupitz and Sebastien Boutet and Hansen, {Debra T.} and Kirian, {Richard A.} and Marius Schmidt and Raimund Fromme and Matthias Frank and Alexandra Ros and Chen, {Julian J.L.} and Sabine Botha and Petra Fromme",

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doi = "10.1016/j.str.2022.12.009",

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AU - Jernigan, Rebecca J.

AU - Logeswaran, Dhenugen

AU - Doppler, Diandra

AU - Nagaratnam, Nirupa

AU - Sonker, Mukul

AU - Yang, Jay How

AU - Ketawala, Gihan

AU - Martin-Garcia, Jose M.

AU - Shelby, Megan L.

AU - Grant, Thomas D.

AU - Mariani, Valerio

AU - Tolstikova, Alexandra

AU - Sheikh, Michelle Z.

AU - Yung, Mimi Cho

AU - Coleman, Matthew A.

AU - Zaare, Sahba

AU - Kaschner, Emily K.

AU - Rabbani, Mohammad Towshif

AU - Nazari, Reza

AU - Zacks, Michele A.

AU - Hayes, Brandon

AU - Sierra, Raymond G.

AU - Hunter, Mark S.

AU - Lisova, Stella

AU - Batyuk, Alexander

AU - Kupitz, Christopher

AU - Boutet, Sebastien

AU - Hansen, Debra T.

AU - Kirian, Richard A.

AU - Schmidt, Marius

AU - Fromme, Raimund

AU - Frank, Matthias

AU - Ros, Alexandra

AU - Chen, Julian J.L.

AU - Botha, Sabine

AU - Fromme, Petra

PY - 2023/2/2

Y1 - 2023/2/2

N2 - NendoU from SARS-CoV-2 is responsible for the virus's ability to evade the innate immune system by cleaving the polyuridine leader sequence of antisense viral RNA. Here we report the room-temperature structure of NendoU, solved by serial femtosecond crystallography at an X-ray free-electron laser to 2.6 Å resolution. The room-temperature structure provides insight into the flexibility, dynamics, and other intrinsic properties of NendoU, with indications that the enzyme functions as an allosteric switch. Functional studies examining cleavage specificity in solution and in crystals support the uridine-purine cleavage preference, and we demonstrate that enzyme activity is fully maintained in crystal form. Optimizing the purification of NendoU and identifying suitable crystallization conditions set the benchmark for future time-resolved serial femtosecond crystallography studies. This could advance the design of antivirals with higher efficacy in treating coronaviral infections, since drugs that block allosteric conformational changes are less prone to drug resistance.

AB - NendoU from SARS-CoV-2 is responsible for the virus's ability to evade the innate immune system by cleaving the polyuridine leader sequence of antisense viral RNA. Here we report the room-temperature structure of NendoU, solved by serial femtosecond crystallography at an X-ray free-electron laser to 2.6 Å resolution. The room-temperature structure provides insight into the flexibility, dynamics, and other intrinsic properties of NendoU, with indications that the enzyme functions as an allosteric switch. Functional studies examining cleavage specificity in solution and in crystals support the uridine-purine cleavage preference, and we demonstrate that enzyme activity is fully maintained in crystal form. Optimizing the purification of NendoU and identifying suitable crystallization conditions set the benchmark for future time-resolved serial femtosecond crystallography studies. This could advance the design of antivirals with higher efficacy in treating coronaviral infections, since drugs that block allosteric conformational changes are less prone to drug resistance.

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JO - Structure

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Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser

Abstract

Keywords

ASJC Scopus subject areas

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Room temperature structure of NSP15 Endoribonuclease from SARS CoV-2 solved using SFX.

Cite this

Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser

Abstract

Keywords

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Datasets

Room temperature structure of NSP15 Endoribonuclease from SARS CoV-2 solved using SFX.

Cite this