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

Journal Contribution - Journal Article

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.
Journal: Acta Crystallographica Section F
ISSN: 2053-230X
Issue: 10
Volume: 77
Pages: 374-384
Number of pages: 11
Publication year:2021
Keywords:Nanobody aided crystallography, Toxin-Antitoxin module, Transcription regulation, Transcription Factor, Structural Biology, Crystallography, Molecular Biophysics