Small-angle X-ray scattering- and nuclear magnetic resonance-derived conformational ensemble of the highly flexible antitoxin PaaA2.
Antitoxins from prokaryotic type II toxin-antitoxin modules are characterized by a high degree of intrinsic disorder. The description of such highly flexible proteins is challenging because they cannot be represented by a single structure. Here, we present a combination of SAXS and NMR data to describe the conformational ensemble of the PaaA2 antitoxin from the human pathogen E. coli O157. The method encompasses the use of SAXS data to filter ensembles out of a pool of conformers generated by a custom NMR structure calculation protocol and the subsequent refinement by a block jackknife procedure. The final ensemble obtained through the method is validated by an established residual dipolar coupling analysis. We show that the conformational ensemble of PaaA2 is highly compact and that the protein exists in solution as two preformed helices, connected by a flexible linker, that probably act as molecular recognition elements for toxin inhibition.
Structural Biology Brussels, Department of Biotechnology, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium; Molecular Recognition Unit and Jean Jeener NMR Centre, Structural Biology Research Center, VIB, Pleinlaan 2, B-1050 Brussels, Belgium.
Yann G J
YG
Sterckx
Structural Biology Brussels, Department of Biotechnology, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium; Molecular Recognition Unit and Jean Jeener NMR Centre, Structural Biology Research Center, VIB, Pleinlaan 2, B-1050 Brussels, Belgium.
Alexander N
AN
Volkov
Structural Biology Brussels, Department of Biotechnology, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium; Molecular Recognition Unit and Jean Jeener NMR Centre, Structural Biology Research Center, VIB, Pleinlaan 2, B-1050 Brussels, Belgium.
Wim F
WF
Vranken
Protein Dynamics and Flexibility, Institut de Biologie Structurale Jean-Pierre Ebel CNRS-CEA-UJF UMR 5075, 41 Rue Jules Horowitz, 38027 Grenoble Cedex, France.
Jaka
J
Kragelj
Protein Dynamics and Flexibility, Institut de Biologie Structurale Jean-Pierre Ebel CNRS-CEA-UJF UMR 5075, 41 Rue Jules Horowitz, 38027 Grenoble Cedex, France.
Malene Ringkjøbing
MR
Jensen
Structural Biology Brussels, Department of Biotechnology, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium; Molecular Recognition Unit and Jean Jeener NMR Centre, Structural Biology Research Center, VIB, Pleinlaan 2, B-1050 Brussels, Belgium.
Lieven
L
Buts
Structural Biology Brussels, Department of Biotechnology, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium; Molecular Recognition Unit and Jean Jeener NMR Centre, Structural Biology Research Center, VIB, Pleinlaan 2, B-1050 Brussels, Belgium.
Abel
A
Garcia-Pino
Laboratoire de Génétique et Physiologie Bactérienne, Institut de Biologie et de Médecine Moléculaires Faculté des Sciences, Université Libre de Bruxelles, 12 Rue des Professeurs Jeener et Brachet, B-6041 Gosselies, Belgium.
Thomas
T
Jové
Laboratoire de Génétique et Physiologie Bactérienne, Institut de Biologie et de Médecine Moléculaires Faculté des Sciences, Université Libre de Bruxelles, 12 Rue des Professeurs Jeener et Brachet, B-6041 Gosselies, Belgium.
Laurence
L
Van Melderen
Protein Dynamics and Flexibility, Institut de Biologie Structurale Jean-Pierre Ebel CNRS-CEA-UJF UMR 5075, 41 Rue Jules Horowitz, 38027 Grenoble Cedex, France.
Martin
M
Blackledge
Structural Biology Brussels, Department of Biotechnology, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium; Molecular Recognition Unit and Jean Jeener NMR Centre, Structural Biology Research Center, VIB, Pleinlaan 2, B-1050 Brussels, Belgium. Electronic address: nico.vannuland@vib-vub.be.
Nico A J
NA
van Nuland
Structural Biology Brussels, Department of Biotechnology, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussels, Belgium; Molecular Recognition Unit and Jean Jeener NMR Centre, Structural Biology Research Center, VIB, Pleinlaan 2, B-1050 Brussels, Belgium. Electronic address: remy.loris@vib-vub.be.
Remy
R
Loris
D000595:Amino Acid Sequence; D000992:Antitoxins / Q000737:chemistry* / Q000235:genetics; D001427:Bacterial Toxins / Q000737:chemistry* / Q000235:genetics; D019453:Escherichia coli O157 / Q000737:chemistry* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D053838:Scattering, Small Angle; D014961:X-Ray Diffraction; D051841:Transcription Factor RelB
D000992:Antitoxins; D001427:Bacterial Toxins; D029968:Escherichia coli Proteins; C520614:Paa protein, E coli; C465567:RelB protein, E coli; D051841:Transcription Factor RelB