Nanobody-aided crystallization of the transcription regulator PaaR2 from Escherichia coli O157:H7

Pieter De Bruyn; Marusa Prolic Kalinsek; Alexandra Vandervelde; Milan Malfait; Yann Sterckx; Frank Sobott; San Hadzi; Els Pardon; Jan Steyaert; Remy Loris

doi:10.1107/S2053230X21009006

Nanobody-aided crystallization of the transcription regulator PaaR2 from Escherichia coli O157:H7

Pieter De Bruyn, Marusa Prolic Kalinsek, Alexandra Vandervelde, Milan Malfait, Yann Sterckx, Frank Sobott, San Hadzi, Els Pardon, Jan Steyaert, Remy Loris

Onderzoeksoutput: Article › peer review

1 Citaat (Scopus)

310 Downloads (Pure)

Samenvatting

paaR2-paaA2-parE2 is a three-component toxin-antitoxin module found in prophage CP-993P of Escherichia coli O157:H7. Transcription regulation of this module occurs via the 123 amino acid regulator PaaR2 which forms a large oligomeric structure. Despite appearing to be well-folded, PaaR2 as well as its N-terminal DNA binding domain withstand crystallization. Native mass spectrometry was used to screen for nanobodies that form a unique complex and stabilize the octameric structure of PaaR2. One such nanobody, Nb33, allowed crystallization of the protein. The resulting crystals belong to space group F432 with a= 317 Å, diffract to 4.0 Å resolution and likely contain four PaaR2 monomers and four nanobody monomers in their asymmetric unit. Crystals of two truncates containing the N-terminal helix-turn-helix domain also interact with Nb33 and the corresponding co-crystals diffract to 1.6 and 1.75 Å resolution.

Originele taal-2	English
Artikelnummer	11
Pagina's (van-tot)	374-384
Aantal pagina's	11
Tijdschrift	Acta Crystallographica Section F - Structural Biology Communications
Volume	77
Nummer van het tijdschrift	10
DOI's	https://doi.org/10.1107/S2053230X21009006
Status	Published - 1 okt. 2021

Toegang tot document

10.1107/S2053230X21009006

74396672Final published version, 1,55 MBLicentie: Unspecified
74396672Final published version, 1,47 MBLicentie: Unspecified

https://eprints.whiterose.ac.uk/180323/6/S2053230X21009006.pdfLicentie: Unspecified

Handle.net

Permanente koppeling

Andere bestanden en links

Link to publication in Scopus

FWOAL831: Regulatie van persistentie door het paaAR-paaA2-parE2 en cog4197-duf1019 operons van E. coli O157
Loris, R. (Administrative Promotor)
1/01/17 → 31/12/20
Project: Fundamenteel

1 Citaties
4 Article
1 PhD Thesis

Bistable expression of a toxin-antitoxin system located in a cryptic prophage of Escherichia coliO157:H7
Jurenas, D., Fraikin, N., Goormaghtigh, F., De Bruyn, P., Vandervelde, A., Jove, T., Charlier, D., Loris, R. & Van Melderen, L., 21 dec. 2021, In: mBio. 12, 6, 15 blz., e0294721.
Onderzoeksoutput: Article › peer review

Open Access
Bestand
16 Citaten (Scopus)

254 Downloads (Pure)
Prokaryote toxin-antitoxin modules: complex regulation of an unclear function
De Bruyn, P., Girardin, Y. & Loris, R., jun. 2021, In: Protein Sci. 30, 6, blz. 1103-1113 11 blz.
Onderzoeksoutput: Article › peer review

Open Access
Bestand
28 Citaten (Scopus)

196 Downloads (Pure)
Toxin-antitoxin modules: functional characterization of transcription regulation and potential use against pathogenic bacteria
De Bruyn, P., 2021, Brussels.
Onderzoeksoutput: PhD Thesis

PaaR2 regulator N-terminal domain
Loris, R. (Project Leader), Volkov, O. (Creator) & Prolic Kalinsek, M. (Creator), Protein Data Bank, 2 mrt. 2022
DOI: 10.2210/pdb7NKV/pdb
Dataset

PhD Defense Mariusa Prolic-Kalinsek - Regulatory interplay between the toxin-antitoxin cluster paaR2-paaA2-parE2 and prophage genes ydaS-ydaT from Escherichia coliO157:H7 (Evenement)
Stijlemans, B. (Jury), Prolic Kalinsek, M. (Speaker), Tompa, P. (Chair), Brunner, J. (Jury), De Proft, F. (Jury), Strelkov, S. (Jury), Bouckaert, J. (Jury) & Loris, R. (Supervisor)
16 mei 2023
Activiteit: Member of PhD committee
PhD Thesis defense Pieter De Bruyn - Toxin-antitoxin modules: functional characterization of transcription regulation and potential use against pathogenic bacteria (Evenement)
Peeters, E. (Chair), Van Molle, I. (Jury), Roelants, K. (Jury), Loris, R. (Supervisor), De Greve, H. (Supervisor), Tenson, T. (Jury) & Michiels, J. (Jury)
2 sep. 2021
Activiteit: Member of PhD committee

Citeer dit

De Bruyn, P., Prolic Kalinsek, M., Vandervelde, A., Malfait, M., Sterckx, Y., Sobott, F., Hadzi, S., Pardon, E., Steyaert, J., & Loris, R. (2021). Nanobody-aided crystallization of the transcription regulator PaaR2 from Escherichia coli O157:H7. Acta Crystallographica Section F - Structural Biology Communications, 77(10), 374-384. Artikel 11. https://doi.org/10.1107/S2053230X21009006

@article{f57fbeb7ea0c4e9f9e7f15753e9a1050,

title = "Nanobody-aided crystallization of the transcription regulator PaaR2 from Escherichia coli O157:H7",

abstract = "paaR2-paaA2-parE2 is a three-component toxin-antitoxin module found in prophage CP-993P of Escherichia coli O157:H7. Transcription regulation of this module occurs via the 123 amino acid regulator PaaR2 which forms a large oligomeric structure. Despite appearing to be well-folded, PaaR2 as well as its N-terminal DNA binding domain withstand crystallization. Native mass spectrometry was used to screen for nanobodies that form a unique complex and stabilize the octameric structure of PaaR2. One such nanobody, Nb33, allowed crystallization of the protein. The resulting crystals belong to space group F432 with a= 317 {\AA}, diffract to 4.0 {\AA} resolution and likely contain four PaaR2 monomers and four nanobody monomers in their asymmetric unit. Crystals of two truncates containing the N-terminal helix-turn-helix domain also interact with Nb33 and the corresponding co-crystals diffract to 1.6 and 1.75 {\AA} resolution.",

keywords = "Nanobody aided crystallography, Toxin-Antitoxin module, Transcription regulation, Transcription Factor, Structural Biology, Crystallography, Molecular Biophysics",

author = "{De Bruyn}, Pieter and {Prolic Kalinsek}, Marusa and Alexandra Vandervelde and Milan Malfait and Yann Sterckx and Frank Sobott and San Hadzi and Els Pardon and Jan Steyaert and Remy Loris",

year = "2021",

month = oct,

day = "1",

doi = "10.1107/S2053230X21009006",

language = "English",

volume = "77",

pages = "374--384",

journal = "Acta Crystallographica Section F - Structural Biology Communications",

issn = "2053-230X",

publisher = "International Union of Crystallography",

number = "10",

}

De Bruyn, P, Prolic Kalinsek, M, Vandervelde, A, Malfait, M, Sterckx, Y, Sobott, F, Hadzi, S, Pardon, E , Steyaert, J & Loris, R 2021, 'Nanobody-aided crystallization of the transcription regulator PaaR2 from Escherichia coli O157:H7', Acta Crystallographica Section F - Structural Biology Communications, vol. 77, nr. 10, 11, blz. 374-384. https://doi.org/10.1107/S2053230X21009006

TY - JOUR

T1 - Nanobody-aided crystallization of the transcription regulator PaaR2 from Escherichia coli O157:H7

AU - De Bruyn, Pieter

AU - Prolic Kalinsek, Marusa

AU - Vandervelde, Alexandra

AU - Malfait, Milan

AU - Sterckx, Yann

AU - Sobott, Frank

AU - Hadzi, San

AU - Pardon, Els

AU - Steyaert, Jan

AU - Loris, Remy

PY - 2021/10/1

Y1 - 2021/10/1

N2 - paaR2-paaA2-parE2 is a three-component toxin-antitoxin module found in prophage CP-993P of Escherichia coli O157:H7. Transcription regulation of this module occurs via the 123 amino acid regulator PaaR2 which forms a large oligomeric structure. Despite appearing to be well-folded, PaaR2 as well as its N-terminal DNA binding domain withstand crystallization. Native mass spectrometry was used to screen for nanobodies that form a unique complex and stabilize the octameric structure of PaaR2. One such nanobody, Nb33, allowed crystallization of the protein. The resulting crystals belong to space group F432 with a= 317 Å, diffract to 4.0 Å resolution and likely contain four PaaR2 monomers and four nanobody monomers in their asymmetric unit. Crystals of two truncates containing the N-terminal helix-turn-helix domain also interact with Nb33 and the corresponding co-crystals diffract to 1.6 and 1.75 Å resolution.

AB - paaR2-paaA2-parE2 is a three-component toxin-antitoxin module found in prophage CP-993P of Escherichia coli O157:H7. Transcription regulation of this module occurs via the 123 amino acid regulator PaaR2 which forms a large oligomeric structure. Despite appearing to be well-folded, PaaR2 as well as its N-terminal DNA binding domain withstand crystallization. Native mass spectrometry was used to screen for nanobodies that form a unique complex and stabilize the octameric structure of PaaR2. One such nanobody, Nb33, allowed crystallization of the protein. The resulting crystals belong to space group F432 with a= 317 Å, diffract to 4.0 Å resolution and likely contain four PaaR2 monomers and four nanobody monomers in their asymmetric unit. Crystals of two truncates containing the N-terminal helix-turn-helix domain also interact with Nb33 and the corresponding co-crystals diffract to 1.6 and 1.75 Å resolution.

KW - Nanobody aided crystallography

KW - Toxin-Antitoxin module

KW - Transcription regulation

KW - Transcription Factor

KW - Structural Biology

KW - Crystallography

KW - Molecular Biophysics

UR - http://www.scopus.com/inward/record.url?scp=85116611532&partnerID=8YFLogxK

U2 - 10.1107/S2053230X21009006

DO - 10.1107/S2053230X21009006

M3 - Article

C2 - 34605442

SN - 2053-230X

VL - 77

SP - 374

EP - 384

JO - Acta Crystallographica Section F - Structural Biology Communications

JF - Acta Crystallographica Section F - Structural Biology Communications

IS - 10

M1 - 11

ER -

Nanobody-aided crystallization of the transcription regulator PaaR2 from Escherichia coli O157:H7

Samenvatting

Toegang tot document

Handle.net

Andere bestanden en links

Vingerafdruk

Projecten

FWOAL831: Regulatie van persistentie door het paaAR-paaA2-parE2 en cog4197-duf1019 operons van E. coli O157

Onderzoekersoutput

Bistable expression of a toxin-antitoxin system located in a cryptic prophage of Escherichia coliO157:H7

Prokaryote toxin-antitoxin modules: complex regulation of an unclear function

Toxin-antitoxin modules: functional characterization of transcription regulation and potential use against pathogenic bacteria

Datasets

PaaR2 regulator N-terminal domain

Activiteiten

PhD Defense Mariusa Prolic-Kalinsek - Regulatory interplay between the toxin-antitoxin cluster paaR2-paaA2-parE2 and prophage genes ydaS-ydaT from Escherichia coliO157:H7 (Evenement)

PhD Thesis defense Pieter De Bruyn - Toxin-antitoxin modules: functional characterization of transcription regulation and potential use against pathogenic bacteria (Evenement)

Citeer dit